Hennara/MedRAG_contents · Datasets at Hugging Face

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f290f0c8-2816-41a1-8f96-399e6d91ff89	2	Nonencapsulated strains are excellent transformation recipients. Glycine-induced pleomorphism reduces the transformation frequency and the degree of inhibition is dependent on the phosphate concentration of the medium. Rifampin resistance and shifts from adenine, hypoxanthine, uracil, and nitrogenase auxotrophy to prototrophy can be achieved. Although single marker transfer is always greater than double marker transfer, the data suggest that rifampin resistance is linked to hypoxanthine, adenine and uracil protorophy at intervals of increasing distance.
f290f0c8-2816-41a1-8f96-399e6d91ff89	3	Rifampin resistance did not appear to be linked to nitrogenase.
48a936ef-68c5-4d35-81b2-c19ac5469298	0	Role of D-tryptophan oxidase in D-tryptophan utilization by Escherichia coli. Mutants of Escherichia coli K-12 that require L-tryptophan (trp) are normally unable to utilize D-tryptophan to fulfill their requirement. However, secondary mutations (dadR) that confer this ability can be isolated. In such strains two distinct enzymes are found to be produced at high levels:
48a936ef-68c5-4d35-81b2-c19ac5469298	1	D-amino acid oxidase (EC 1.4.3.3) and D-tryptophan oxidase. A convenient assay procedure for D-tryptophan oxidase is described. The two enzymes could be distinguished on the basis of their sensitivity to inhibition by L-phenylalanine and L-tyrosine. Strains that were trp dadR could not grow with D-tryptophan in the presence of L-phenylalanine, but further mutations, Fyo, could be isolated that allowed growth under these conditions.
48a936ef-68c5-4d35-81b2-c19ac5469298	2	Some of them were characterized by further increases in the level of D-tryptophan oxidase activity and a sharp decrease in D-amino acid oxidase. These kinds of Fyo mutations lay in or near the dadR gene. The substrate specificity of the two enzymes toward a large number of compounds was examined.
48a936ef-68c5-4d35-81b2-c19ac5469298	3	The transamination of aromatic keto acids was investigated. In the wild-type strain only a single enzyme, transaminase A (EC 2.6.1.5), was found, and it was irreversibly activated when subjected to elevated temperatures. The present state of our knowledge on D-amino acid utilization in E.
48a936ef-68c5-4d35-81b2-c19ac5469298	4	coli is summarized.
74c3405b-1582-4c08-aa62-9679284c4b0c	0	Transduction of chromosomal genes between enteric bacteria by bacteriophage P1. We have used P1 transduction to create intergeneric hybrid strains of enteric bacteria by moving the genA and hut genes between Klebsiella aerogenes, Escherichia coli and Salmonella typhimurium. The use of E. coli as the recipient in such transductions permits the construction of episomes and specialized transducing phage containing non-E. coli material. The effect of host restriction modification and deoxyribonucleic acid homology on the frequency of intergeneric transduction of these loci has been examined.
a9eede7e-25ba-4c7c-a675-4875b88c56a7	0	Galactoside accumulation by Escherichia coli, driven by a pH gradient. Acidification of the external medium results in thiomethylgalactoside accumulation in an energy-depleted adenosine triphosphatase-negative mutant of Escherichia coli.
db033684-25ab-4037-87e2-aa0a6393beef	0	Adenosine 5'-monophosphate-stimulated cyanide-insensitive respiration in mitochondria of Moniliella tomentosa. Mitochondria of the yeastlike fungus Moniliella tomentosa oxidize reduced nicotinamide adenine dinucleotide, reduced nicotinamide adenine dinucleotide phosphate, succinate, isocitrate, and lactate. These oxidations are completely inhibited by cyanide or antimycin A in mitochondria isolated from cells grown in the standard medium.
db033684-25ab-4037-87e2-aa0a6393beef	1	On the other hand, the oxidation of all substrates, except lactate, is almost completely insensitive to cyanide or antimycin A in mitochondria from cells grown in the presence of ethidium bromide. In this instance, the oxidation is mainly mediated by an alternate oxidase which can be blocked by salicyl hydroxamic acid.
db033684-25ab-4037-87e2-aa0a6393beef	2	The alternate oxidase can be specifically stimulated by adenosine 5'-monophosphate and this provides a new method for the characterization of the alternate oxidase in mitochondria of M. tomentosa.
1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e	0	Sulfate-reducing pathway in Escherichia coli involving bound intermediates. Although a sulfate-reducing pathway in Escherichia coli involving free sulfite and sulfide has been suggested, it is shown that, as in Chlorella, a pathway involving bound intermediates is also present. E. coli extracts contained a sulfotransferase that transferred the sulfonyl group from a nucleosidephosphosulfate to an acceptor to form an organic thiosulfate.
1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e	1	This enzyme was specific for adenosine 3'-phosphate 5'-phosphosulfate, did not utilize adenine 5'-phosphosulfate, and transferred to a carrier molecule that was identical with thioredoxin in molecular weight and amino acid composition. In the absence of thioredoxin, only very low levels of the transfer of the sulfo group to thiols was observed.
1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e	2	As in Chlorella, thiosulfonate reductase activity that reduced glutathione-S-SO3- to bound sulfide could be detected. In E. coli, this enzyme used reduced nicotinamide adenine dinucleotide phosphate and Mg2+, but did not require the addition of ferredoxin or ferredoxin nicotinamide adenine dinucleotide phosphate reductase. Although in Chlorella the thiosulfonate reductase appears to be a different enzyme from the sulfite reductase, the E.
1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e	3	coli thiosulfonate reductase and sulfite reductase may be activities of the same enzyme.
dd366adb-2b4f-4d95-a0f5-ba82d756ff7f	0	An endonuclease from Escherichia coli that introduces single polynucleotide chain scissions in ultraviolet-irradiated DNA. An endonuclease that makes single polynucleotide chain scissions in ultraviolet-irradiated DNA has been purified from Escherichia coli. The activity has the following properties: (a) unirradiated DNA is attacked very little if at all;
dd366adb-2b4f-4d95-a0f5-ba82d756ff7f	1	(b) single strand DNA is not attacked, whether irradiated or not; (c) there is no requirement for divalent cations and the activity is not affected by the addition of EDTA; (d) the pH optimum is approximately 7; (e) the activity is inhibited by 1 M NaCl, single strand DNA, transfer RNA and double strand DNA;
dd366adb-2b4f-4d95-a0f5-ba82d756ff7f	2	(f) the sedimentation coefficient, S20,w, is approximately 2.6; (g) it is a basic protein. The enzyme is tentatively named E. coli endonuclease III. The physiological function of the endonuclease has not yet been established.
4269d979-ab90-4056-b518-4eced3f4b7dd	0	Quaternary conformational changes in human hemoglobin studied by laser photolysis of carboxyhemoglobin. These experiments indicate that absorbance changes observed at the 425 nm isosbestic point of the Hb and HbCO following laser photolysis of HbCO provide a direct measure of the rates of quaternary conformational changes between rapidly reacting Hb (the immediate product of full photolysis) and slowly reacting normal deoxyhemoglobin.
4269d979-ab90-4056-b518-4eced3f4b7dd	1	Hb, first observed by Gibson (Gibson, Q.H. (1959) Biochem. J. 71, 293-303), Has been interpreted as deoxyhemoglobin remaining in the liganded quaternary conformation following rapid removal of ligand by a light pulse. In borate buffers between pH 8.4 and 9.6 particularly simple pH-independent results were obtained which allowed the use of a Monod.
4269d979-ab90-4056-b518-4eced3f4b7dd	2	Wyman, and Changeux model (Monod, J., Wyman, J., and Changeux, J (1965) J. Mol. Biol. 12, 88-118) to fit the data. In this case Hb is taken to be R state deoxyhemoglobin. Partial photolysis experiments at 425 nm show that the rate of the R - T conformational change at 20 degrees decreases by about a factor of 2 for each additional bound ligand.
4269d979-ab90-4056-b518-4eced3f4b7dd	3	The rate of the ligand-free conformational change is found to be 920 +/- 60s(-1), 6400 +/- 600s(-1), and 15,700 +/- 700(-1) respectively at 3 degrees, 20 degrees, and 30 degrees. The previously uninterpreted effects of flash length and partial photolysis on the CO recombination kinetics can be explained in terms of the present model.
4269d979-ab90-4056-b518-4eced3f4b7dd	4	Kinetic results obtained below pH 8 are found to be inconsistent with a two-state model. It appears that binding of inositol hexaphosphate produces a new rapidly reacting quaternary conformation of HbCO.
ecfd0bd1-87f5-447c-ae0d-1e2815db0c9b	0	Muscarinic acetylcholine receptor from rat brain. Partial purification and characterization. A protein capable of binding atropine and (3H)propylbenzilylcholine mustard was solubilized and purified (200-fold) from rat brain. Pronase and trypsin, but not phospholipases, diminished the binding capacity of the solubilized receptor. The molecular weight of the salt-solubilized receptor as determined by gel filtration in the absence of detergents is 30,000.
ecfd0bd1-87f5-447c-ae0d-1e2815db0c9b	1	The purified protein showed specificity of binding toward muscarinic ligands. the high and low affinity dissociation constants of the receptor.atropine complex are 0.3 nM and 0.15 muM. Binding of atropine is pH-dependent with an optimum at 7.1. Ca2+ influences the binding of atropine and maximal binding occurs at 0.5
ecfd0bd1-87f5-447c-ae0d-1e2815db0c9b	2	mM Ca2+. The subcellular distribution of the receptor was also examined.
5e00ac47-ef47-4711-9088-6c6887b5b416	0	Quantitative relationships between phosphorylation, electron flow, and internal hydrogen ion concentrations in spinach chloroplasts. 1. Further evidence that the uptake of [14C]hexylamine, determined by centrifugal filtration of spinach chloroplast thylakoids through silicone fluid layers, gives precise estimations of light-induced H+ concentration gradients (deltapH) is presented.
5e00ac47-ef47-4711-9088-6c6887b5b416	1	DeltapH was independent of the amount of thylakoids used or of the concentration of hexylamine. Moreover, hexylamine uptake was sensitive to the osmolarity of the suspending medium. 2. Internal H+ concentration ([H+]in) is proportional to the rate of electron flow when light intensity was used to vary these parameters.
5e00ac47-ef47-4711-9088-6c6887b5b416	2	Proportionality was still observed in the presence of 0.1 and 1.0 muM gramicidin D. When, however, [H+]in and electron flow were varied by increasing the concentration of gramicidin D, at constant light intensity the rate of electron flow was approximately proportional to 1/[H]in.
5e00ac47-ef47-4711-9088-6c6887b5b416	3	3. The phosphorylation efficiency (P/e2 ratio) falls with decreasing light intensity or increasing concentrations of the phosphorylation inhibitor, 4'-deoxyphlorizin. The proportionality between the rate of electron flow and [H+]in allows the calculation of the rate of nonphosphorylating (basal) electron flow if [H+]in under phosphorylating conditions is known.
5e00ac47-ef47-4711-9088-6c6887b5b416	4	The contribution of basal electron flow, a consequence of passive efflux of H+ from the thylakoids, to the overall rate of electron flow increases as the rate of phosphorylation decreases. P/e2 ratios calculated using rates of electron flow from which the basal component has been subtracted are constant.
5e00ac47-ef47-4711-9088-6c6887b5b416	5	A calculated P/e2 ratio of about 1.3 is obtained. 4. It is shown that the reciprocal of the phosphorylation efficiency should be proportional to 1/[H+]in2 when these parameters are varied using light intensity. This relationship was verified and provided an estimate of the P/e2 at infinite [H+]in.
5e00ac47-ef47-4711-9088-6c6887b5b416	6	This value was 1.3. These results provide further evidence that a H+ electrochemical gradient serves to couple photophosphorylation to electron flow and that the rate of phosphorylation is proportional to [H+]in3. That is, three H+ are translocated out of thylakoids for each adenosine triphosphate formed.
c34b4ecf-680c-497c-9ff0-fad105f6120b	0	Kinetic properties of human placental aromatase. Application of an assay measuring 3H2O release from 1beta,2beta-3H-androgens. The rapid and sensitive assay of 1beta,2beta-3H-androgen aromatization by measurement of 3H2O release (Thompson, E.A., Jr., and Siiteri, P.K. (1974) J. Biol.
c34b4ecf-680c-497c-9ff0-fad105f6120b	1	Chem. 249, 5364-5372) has been analyzed to determine its applicability to initial rate studies. It was found that aromatization is the sole reaction catalyzed by lyophilized placental microsomes that causes a loss of tritium from position 1 or 2 of androstenedione and testosterone. Tritium is, however, removed from position 2 of the estrogen products, presumably in 2-hydroxylation, but this does not invalidate use of the assay for initial rate measurements;
c34b4ecf-680c-497c-9ff0-fad105f6120b	2	it was therefore used to characterize the catalytic properties of aromatase. Aromatization by the freeze-dried preparation was stimulated by K+, EDTA, and dithiothreitol, and was maximally active at pH 7.5 TO 8.0. With incubation conditions optimized for these factors, the apparent Km for NADPH is approximately 1 muM.
c34b4ecf-680c-497c-9ff0-fad105f6120b	3	The maximum velocity of androstenedione aromatization exceeds that of testosterone, and the affinity of the substrate binding site is higher for the former substrate, the apparent Km values being 0.1 muM and 0.4 muM, respectively. Mutual competition experiments with the androgen substrates showed that each gives simple competitive inhibition of the other's aromatization;
c34b4ecf-680c-497c-9ff0-fad105f6120b	4	furthermore, the apparent Ki values for each are in close agreement with their respective Km values. Androst-1,4,6-triene-3,17-dione competitively inhibits the aromatization of both androstenedione and testosterone, the apparent Ki, in both cases being 0.2 muM. It is concluded that the two androgen substrates are aromatized at a single, identical site.
9471d7ca-281e-466f-957f-5f8ef72b4aca	0	Apparent oxidation-reduction potential of Clostridium acidi-urici ferredoxin. Effect of pH, ionic strength, and amino acid replacements. The effects of pH and ionic strength on the midpoint reduction potential (Emp) of Clostridium acidi-urici ferredoxin were determined using hydrogen gas and hydrogenase. The Emp of native ferredoxin at 24-25 degrees in 0.1
9471d7ca-281e-466f-957f-5f8ef72b4aca	1	M Tris-chloride buffer, pH 7.0, is--0.434 V. In the pH range examined, the Emp becomes approximately 13 mv more negative per each pH unit increase. A plot of the log of ionic strength versus the apparent Emp of ferredoxin in 0.1 M Tris-chloride buffer, pH 7.5
9471d7ca-281e-466f-957f-5f8ef72b4aca	2	, Was linear over the range of 1.0 to 0.01 ionic strength with Emp values of--0.414 and--0.475 V, respectively, at these extremes. This effect is the same with sodium chloride, sodium bromide, or ammonium sulfate. Potassium phosphate buffer caused a similar change, but the absolute values of Emp differed from those obtained in the presence of the other salts.
9471d7ca-281e-466f-957f-5f8ef72b4aca	3	This effect of pH and ionic strength on Emp may be general for clostridial-type (Fe4S4)2-ferredoxins, since the apparent Emp of Clostridium pasteurianum ferredoxin is affected in a similar manner by these two variables. The Emp of this ferredoxin in 0.1 M Tris-chloride buffer pH 7.0
9471d7ca-281e-466f-957f-5f8ef72b4aca	4	, is--0.405 V. Since the NH2-terminal amino acid residue, Ala1, and Tyr2 of C. acidi urici ferredoxin are near an (Fe4S4)2-cluster in the protein, the apparent Emp of derivatives that contained amino acid replacements in these two positions were determined. Under similar conditions, the Emp of most of the 13 derivatives examined, including those of [Leu2]- and[3-NH2-Tyr30]ferredoxin, is approximately the same as that of native ferredoxin.
9471d7ca-281e-466f-957f-5f8ef72b4aca	5	However, the Emp of [His2]ferredoxin is approximately 15 mv more positive, whereas that of [Trp2]ferredoxin is 22 mv more negative than that of native C. acidi-urici ferredoxin. Variations in sodium chloride concentration and pH also affected the apparent Emp of the derivatives. It is suggested that the changes observed in the Emp of C.
9471d7ca-281e-466f-957f-5f8ef72b4aca	6	acidi-urici ferredoxin are caused by protein conformational changes.
c42335f1-02b8-471d-ad22-742cf535d1ba	0	A deoxyribonucleic acid kinase from nuclei of rat liver. Purification and properties. A DNA kinase has been partially purified from rat liver nuclei by a procedure which also yields DNA ligase. The kinase uses ATP to phosphorylate specifically the 5'-hydroxyl termini of oligodeoxynucleotides and of single- or double-stranded DNA, yielding 5'-phosphate termini and ADP.
c42335f1-02b8-471d-ad22-742cf535d1ba	1	The kinase is inactive on RNA, or on oligodeoxynucleotides of chain length less than approximately 10 to 12 residues. The kinase requires a divalent cation (Mg2+, Mn2+, Co2+, Zn2+, Ni2+, or Ca2+) for activity and has an acidic pH optimum. It is inhibited by a variety of nucleotides as well as by very low levels of inorganic and organic sulfate compounds and sulfate analogues.
c42335f1-02b8-471d-ad22-742cf535d1ba	2	The molecular weight of the kinase is estimated to be 8 times 10(4) from gel filtration.
e7c1efed-2035-48a9-b97f-bd8cdd33e243	0	Hydrogen exchange at the amide group of reduced pyridine nucleotides and the inhibition of that reaction by dehydrogenases. Stopped flow ultraviolet spectroscopy has been used to measure the rate of hydrogen exchange with solvent at the amide group of reduced nicotinamide nucleotide coenzymes. Several mechanisms for the exchange reaction are considered in the light of the kinetic data. Complex formation between the coenzyme and any of four dehydrogenases markedly slows the rate of hydrogen exchange. Hydrogen bond formation and/or hydrophobic interactions within these complexes are thought to be the reasons for the decreased rate of exchange.
f2437d01-97be-470f-afc0-2ca92f82b5ff	0	A Dacron wool packed-bed extracorporeal reactor: a kinetic study of immobilized Escherichia coli II L-asparaginase. An extracorporeal reactor containing a packed bed of Dacron fibers has been developed. Escherichia coli II L-asparaginase was coupled to the Dacron using gamma-aminopropyltriethoxysilane and glutaraldehyde. The preparation had an activity of 37 IU per gram of Dacron (37 degrees C).
f2437d01-97be-470f-afc0-2ca92f82b5ff	1	The apparent Km was studied as a function of the flow rate. The data indicated that the apparent Km approached the Km of the native enzyme at flow rates of about 300 mg/min. In vivo use of L-asparaginase immobilized on the Dacron indicated effective lowering of plasmatic L-asparagine levels.
653d2110-ff66-465a-b3ec-a0940565fa06	0	Isolation of intact megakaryocytes from guinea pig femoral marrow. Successful harvest made possible with inhibitions of platelet aggregation; enrichment achieved with a two-step separation technique. Methods have been devised to harvest megakaryocytes from guinea pig femoral marrow and to isolate them in high yield. When marrow tissue was disaggregated the megakaryocytes underwent degenerative changes characterized by the loss of cytoplasmic granules and alterations in membrane topography, similar to the changes seen in aggregating platelets.
653d2110-ff66-465a-b3ec-a0940565fa06	1	These morphologic changes were interpreted to mean that megakaryocytes possessed functional attributes of platelets. The use of agents which inhibit platelt aggregation (0.38% sodium citrate. 10(-3) M adenosine, and 2 x 10(-3) M theophylline) in a medium free of bivalent cations prevented these changes.
653d2110-ff66-465a-b3ec-a0940565fa06	2	This solution resulted in both an excellent morphologic preservation and a significantly increased recovery of megakaryocytes from marrow tissue. A two-step purification of the intact megakaryocytes was carried out on the basis of their low density and large size, with equilibrium density gradient centrifugation followed by velocity sedimentation.
653d2110-ff66-465a-b3ec-a0940565fa06	3	This sequence gave approximately a 100-fold enrichment of megakaryocytes, significantly better than that achieved with either method alone. These techniques for harvesting and concentrating megakaryocytes make it possible for the first time to study megakaryocytes in vitro.
b619ca86-18fe-4f4d-85c9-1250584d54e1	0	The polymerization of actin. III. Aggregates of nonfilamentous actin and its associated proteins: a storage form of actin. When echinoderm sperm are treated with the detergent Triton X-100 at pH 6.4 in 10 mM phosphate buffer, the membranes are solubilized, but the actin which is located in the periacrosomal region remains as a phase-dense cup.
b619ca86-18fe-4f4d-85c9-1250584d54e1	1	These cups can be isolated free from the flagella and chromatin and can be solubilized by increasing the pH to 8.0 and by changing the ionic strength and type of buffer used. Since the actin does not exist in the "F" state in unreacted sperm, and since the actin remains as a unit that does not diffuse away, it must be present in the mature sperm in a bound or storage state.
b619ca86-18fe-4f4d-85c9-1250584d54e1	2	The actin is, in fact, associated with a pair of proteins whose mol wt are 250,000 and 230,000. When the isolated cups are digested with trypsin, these high molecular weight proteins are digested, thereby liberating the actin. The actin will polymerize if heavy meromyosin or subfragment 1 is added to a preparation of isolated cups.
b619ca86-18fe-4f4d-85c9-1250584d54e1	3	Evidence is presented that this pair of high molecular weight proteins is similar in molecular weight and properties to erythrocyte spectrin. Attempts at transforming the storage form of actin in the cup into filaments were only moderately successful. The best conditions for filament formation involve incubating the cup in ATP and divalent salts.
b619ca86-18fe-4f4d-85c9-1250584d54e1	4	Careful examination of these cups reveals that the actin polymerized preferentially on either end of oriented filaments that already exist in the cup, indicating that self-nucleation is inefficacious. I conclude that the actin can exist in the storage form by its association with spectrin-like molecules and that the actin in this state polymerizes preferentially onto existing filaments.
8c62887e-22a7-4bad-8764-aa1f00cedba6	0	Degradation of abnormal proteins in HeLa cells. The experiments show that abnoramal proteins are degraded faster than normal ones in HeLa cells. Among the fragmentary proteins made in the presence of puromycin, those with low molecular weight are least stable. Proteins made after incubation with 5-fluorouracil or in the presence of some amino acid analogues are also unstable. Breakdown of proteins made in the presence or absence of puromycin is nearly unaffected by cycloheximide and is independent of pH between 7 and 8.
44905afb-5ef8-48c7-8d10-5a09806bf399	0	The effect of pH on incorporation of galactose by a normal human cell line and cell lines from patients with defective galactose metabolism. Incorporation of radioactive galactose into TCA-insoluble material of galactosemic fibroblasts is more sensitive to low pH than is the incorporation by normal human fibroblasts.
44905afb-5ef8-48c7-8d10-5a09806bf399	1	This study was undertaken to determine (1) whether there was any pH which could correct or counteract the galactosemic defect relative to galactose incorporation, and (2) whether the low pH effect was specific for galactose metabolism or whether general cellular metabolism in galactosemic cells was more sensitive to low pH than that in normal cells. The pH dependencies of incorporation of radioactive galactose and glucose into cellular macromolecules were investigated in galactosemic and normal cells.
44905afb-5ef8-48c7-8d10-5a09806bf399	2	Normal cells have a biphasic curve with respect to galactose incorporation with peaks at pH 7.0 and 8.5. Galactosemic cells have only the high pH peak. The maximum incorporation by galactosemic cells was never more than about 30% that seen by normal cells under the conditions of these experiments.
44905afb-5ef8-48c7-8d10-5a09806bf399	3	Thus manipulation of the pH alone cannot correct the galactosemic defect. The rate of incorporation of radioactive galactose was studied in normal, galactosemic and galactokinase deficient cells, at pH 7.2 and at pH 6.3. At pH 7.2, galactosemic cells incorporate galactose at a linear rate which is 30 to 40% that of normal cells while incorporation by kinase-deficient cells is between 5 and 10% of normal.
44905afb-5ef8-48c7-8d10-5a09806bf399	4	At pH 6.3, the incorporation is also linear. However, galactosemic cells now exhibit the same rate as kinase-deficient cells in which the low level of incorporation is unaffected by pH. These results suggest that incorporation of galactose by galactosemic cells at low pH is not due to metabolic death of the cells, but may be due to the inhibition of some specific step or steps along a metabolic route of galactose metabolism other than the Leloir pathway.
ce1b057b-8304-41d7-9ae3-869260b84d02	0	Chromatography of hemoglobins on CM-cellulose with bis-tris and sodium chloride developers. CM-Cellulose as an ion-exchange medium with Bis-tris as buffer and a gradient of sodium chloride provides a versatile system for the chromatography of hemoglobins. Changes in pH, Bis-tris concentration, and slope of the sodium chloride grandient provide means for markedly altering chromatographic behavior for special separations. Examples are given of the application of the method to normal samples and to those with hemoglobinopathies.
110fcb13-2778-42f6-9e92-d0b32261ae35	0	The determination of phanquone in biological material by gas-liquid chromatography. A gas-liquid chromatographic method for the quantitative determination of phanquone is described, based on the formation of a dimethoxine prior to its extraction from biological material. The sensitivity of the procedure is about 15 ng/ml in biological fluid.
299ae760-af48-4113-91f6-35053fe45eb6	0	Stability of fluorescent antibody conjugates stored under various conditions. Two experiments were carried out to determine the stability of fluorescent antibody conjugates. In experiment 1, Francisella tularemia conjugates in the lyophilized state retained their original staining titer for 1,294 days when stored at 25, 4 to 5, and -20 C;
299ae760-af48-4113-91f6-35053fe45eb6	1	at 37 C the conjugates were stable for at least 65 days. In the liquid state at pH 7.4 and 8.0 these conjugates were stable for 1,294 days at 4 to 5 and -20 C, whereas those stored at 25 C remained stable through days 473 and 160 of storage, respectively, after which the staining titer gradually dropped.
299ae760-af48-4113-91f6-35053fe45eb6	2	In experiment 2 five previously lyophilized conjugates were rehydrated with three different diluents and stored at 4 to 5 C for up to 600 days at their working dilutions. All of these conjugates retained their original staining titer during the test period except an anti-human globulin conjugate rehydrated with phosphate-buffered saline.
299ae760-af48-4113-91f6-35053fe45eb6	3	Recommendations are made for the long-term storage of fluorescent antibody conjugates.
22dcea31-f492-4856-bcd1-c87458fcc1d0	0	Inactivation of viruses in serum with binary ethyleneimine. The inactivation os six strains from three different groups of viruses with 0.001 M binary ethyleneimine at 37 C proceeded at the same rate in either bovine serum or cell culture medium. The inactivant did not impair the growth-promoting capacity of bovine serum used in cell culture, nor did it affect the antibody activity of guinea pig hyperimmune serum.
c457bed7-c03a-4af8-89bf-d92bbdca4d44	0	Inorganic phosphate homeostasis. Renal adaptation to the dietary intake in intact and thyroparathyroidectomized rats. The possibility of renal tubular adaptation to variations in dietary inorganic phosphate (Pi) was investigated in intact and thyroparathyroidectomized (TPTX) rats pair-fed diets containing low, normal, and high amounts of Pi for periods up to 10 days.
c457bed7-c03a-4af8-89bf-d92bbdca4d44	1	Clearances were measured before and during active i.v. infusions with Pi in conscious animals. Thus tubular reabsorption of phosphate (TRPi) could be assessed over a wide range of plasma phosphate concentrations ([Pi]P1). It was found that the renal tubule could adapt its capacity to transport Pi according to the dietary Pi:
c457bed7-c03a-4af8-89bf-d92bbdca4d44	2	TRPi was always higher, for a given [Pi]P1, in the animals fed low than in those fed higher Pi diets. This diet-induced modification also occurred in the absence of thyroparathyroid glands, in the presence of the same calcemia and urinary pH, and during marked extracellular volume expansion.
c457bed7-c03a-4af8-89bf-d92bbdca4d44	3	A time-course study in rats TPTX both before and during the administration of the experimental diets showed that a difference in the tubular handling of Pi was detectable as early as 3 days after switching the animals from a normal to low- or high-Pi diets. These results indicate that factors other than parathyroid hormone are implicated in the tubular response to variations in the dietary intake of inorganic phosphate.
247316a1-e4e7-4395-8a87-e3ef37de5923	0	Effect of beta adrenergic blockade on renin response to renal nerve stimulation. The ability of d,l-propranolol to block renin secretion in response to various extrarenal stimuli, such as hemorrhage and hypoglycemia, has been interpreted to indicate the presence of an intrarenal beta receptor regulating renin release. However, two problems complicate this interpretation:
247316a1-e4e7-4395-8a87-e3ef37de5923	1	(a) the stimuli have effects outside the kidney, and (b) d,l-propranolol has a local anesthetic, as well as a beta adrenergic blocking, action. In the present study, the effects of a purely intrarenal stimulus, in the form of renal nerve stimulation (RNS), on renin secretion was examined.
247316a1-e4e7-4395-8a87-e3ef37de5923	2	The effects of d,l-propranolol (anesthetic and beta-blocking activity), l-propranolol (beta-blocking activity only), and d-propranolol (local anesthetic activity only) on the renin response to RNS were examined. In a control group of animals, two sequential RNS increased mean renin secretion from 401 to 1,255 U/min (P less than 0.25
247316a1-e4e7-4395-8a87-e3ef37de5923	3	) and from 220 to 2,179 U/min (P less than 0.01). In a second group the first RNS increased renin secretion from 201 to 1,181 U/min (P less than 0.01), but after d,l-propranolol was given RNS did not significantly alter renin secretion (33 to 55 U/min).
247316a1-e4e7-4395-8a87-e3ef37de5923	4	In a third group the initial RNS increased renin secretion from 378 to 1,802 U/min (P less than 0.025), but after l-propranolol was given RNS had no significant effect on renin secretion (84 to 51 U/min). A fourth group of dogs showed a rise in renin secretion from 205 to 880 U/min (P less than 0.001
247316a1-e4e7-4395-8a87-e3ef37de5923	5	) in response to the first RNS, while the second RNS, given after an infusion of d-propranolol, caused a rise in renin secretion from 80 to 482 (P less than 0.005). The nature of the electrical stimulus was consistent in all groups and caused no detectable changes in renal or systemic hemodynamics or in urinary electrolyte excretion.
247316a1-e4e7-4395-8a87-e3ef37de5923	6	The results, therefore, indicate that renin secretion can be stimulated through intrarenal beta receptors independent of changes in systemic or renal hemodynamics or in tubular sodium reabsorption. Hence the effect of beta stimulation on renin secretion would appear to result from a direct action on the renin-secreting cells of the juxtaglomerular apparatus.
de35daee-f381-4e92-b064-8f0f1602076e	0	Identification and characterization of a bile acid receptor in isolated liver surface membranes. It is generally assumed that hepatic transport of bile acids is a carrier-mediated process. However, the basic mechanisms by which these organic anions are translocated across the liver cell surface membrane are not well understood.
de35daee-f381-4e92-b064-8f0f1602076e	1	Since carrier-mediated transport involved binding of the transported molecule to specific receptor sites, we have investigated the possibility that bile acid receptors are present in liver surface membranes. Isolated liver surface membranes were incubated at 4 degrees C with [14C]cholic acid and [14C]taurocholic acid, and membrane-boudn bile acid was separated from free by a rapid ultrafiltration technique through glass-fiber filters.
de35daee-f381-4e92-b064-8f0f1602076e	2	Specific bile acid binding is rapid and reversible and represents approximately 80% of the total bile acid bound to liver surface membranes. Taurocholic acid binding is independent of the medium pH, while cholic acid binding demonstrates an optimum at pH 6.0. Analysis of equilibrium data for both cholic and taurocholic acid binding indicates that specific binding is saturable and consistent with Michaelis-Menten kinetics, while nonspecific binding is nonsaturable.
de35daee-f381-4e92-b064-8f0f1602076e	3	Apparent maximal binding capacity and dissociation constant values indicate a large capacity system of receptors that have an affinity for bile acids comparable to that of the hepatic transport mechanism. Scatchard analysis of the saturation kinetics as well as inhibition studies suggest that bile acids bind to a single and noninteracting class of anion that competes with bile acids for hepatic uptake, also inhibits cholic acid binding.
de35daee-f381-4e92-b064-8f0f1602076e	4	In contrast, no inhibition was demonstrated with indocyanine green and probenecid. Specific bile acid binding is enriched and primarily located in liver surface membranes and found only in tissues involved in bile acid transport. Specific bile acid binding is independnet of Na+, Ca2+, and Mg2+ and does not require metabolic energy.
de35daee-f381-4e92-b064-8f0f1602076e	5	In addition, thiol groups and disulfide are not required for activity at the binding site. However, specific bile acid binding is markedly decreased by low concentrations of proteolytic enzymes and is also decreased by the action of neuraminidase and phospholipases A and C. These results are consistent with the existence of a homogeneous bile acid receptor protein in liver surface membranes.
de35daee-f381-4e92-b064-8f0f1602076e	6	The primary surface membrane location of this receptor, its binding properties, and its ligand specificity suggest that bile acid binding to this receptor may represent the initial interaction in bile acid transport across liver surface membranes.
20ba40d2-3788-40e2-8cce-d41eaca4b6ce	0	A brief anxiety rating scale in evaluating anxiolytics. The Wang Anxiety Rating Scale (WARS) was designed to evaluate degrees of anxiety in patients receiving anxiolytic medication. WARS contains 12 pertinent symptoms of anxiety: nervousness, restlessness, excitability, irritability, worrying, disturbed concentration, palpitation, insomnia, hostility, tremors, smoking, and excessive perspiration.
20ba40d2-3788-40e2-8cce-d41eaca4b6ce	1	Frequently encountered side effects of anxiolytic medications are excluded. The validity of the WARS was determined by correlation with the Hamilton Anxiety Rating Scale (HARS) in a single-blind study in which 20 chronically anxious patients consecutively received placebo (three days), 15 mg clorazepate dipotassium (two weeks), and 22.5
20ba40d2-3788-40e2-8cce-d41eaca4b6ce	2	mg clorazepate dipotassium (two weeks). Both anxiety scales, a side effect scale, and a global assessment were completed at regular intervals (periods 0-6). Results show (1) highly significant correlation (P less than 0.001) between WARS and HARS for periods 1-6; (2) greater correlation between HARS and side effect scale than between WARS and side effect scale;
20ba40d2-3788-40e2-8cce-d41eaca4b6ce	3	(3) greater correlation between WARS and global assessment than between HARS and global assessment; correlated changes in scores for WARS, HARS, and global assessment demonstrate efficacy of active medication.
81bd9e54-081c-481e-b2bd-329c5db8fcdb	0	The changing pattern of bacterial sepsis since the introduction of antibiotic therapy. During the six-year period, 1968-1973, sepsis developed in 1 of every 80 patients admitted to the Presbyterian Hospital, New York. In 1 of 133 patients the sepsis was due to Gram-positive organisms, and in 1 of 188 patients to Gram-negative organisms.
81bd9e54-081c-481e-b2bd-329c5db8fcdb	1	The mortality rate for Gram-positive cases was 4.4 percent, for Gram-negative cases 19.1 percent, and for urologic cases 15.3 percent (versus 56.25 percent in 1959-1964). Data are presented on the relative incidences of involved pathogens in 1740 cases of Gram-positive sepsis /78 deaths), and in 1236 cases of Gram-negative sepsis (205 deaths).
81bd9e54-081c-481e-b2bd-329c5db8fcdb	2	The lowering of the sepsis mortality rate has been the result of preventative measures, early diagnosis, and vigorous treatment. Treatment includes the correction of acidosis and anoxia, early administration of bactericidal antibiotics, and restoration of the microcirculation by administration of corticosteroids, beta-adrenergic drugs, and appropriate diuretics.
0aa0e262-15b9-42ce-bdca-431cd8facce6	0	[Has the pH meter replaced the Apgar score?]. The introduction of measuring the pH appears to place it in competition with Apgar scoring because of its precision. A study of this which has been carried out has illustrated that there are two different criteria for assessing the state of the infant at birth. The usual agreement between pH values and Apgar scoring can be broken when clinical fetal distress has become established before metabolic equilibrium of the infant has become modified. In these circumstances the Apgar score will be bad while the pH will be good.

f290f0c8-2816-41a1-8f96-399e6d91ff89

2

Nonencapsulated strains are excellent transformation recipients. Glycine-induced pleomorphism reduces the transformation frequency and the degree of inhibition is dependent on the phosphate concentration of the medium. Rifampin resistance and shifts from adenine, hypoxanthine, uracil, and nitrogenase auxotrophy to prototrophy can be achieved. Although single marker transfer is always greater than double marker transfer, the data suggest that rifampin resistance is linked to hypoxanthine, adenine and uracil protorophy at intervals of increasing distance.

f290f0c8-2816-41a1-8f96-399e6d91ff89

3

Rifampin resistance did not appear to be linked to nitrogenase.

48a936ef-68c5-4d35-81b2-c19ac5469298

0

Role of D-tryptophan oxidase in D-tryptophan utilization by Escherichia coli. Mutants of Escherichia coli K-12 that require L-tryptophan (trp) are normally unable to utilize D-tryptophan to fulfill their requirement. However, secondary mutations (dadR) that confer this ability can be isolated. In such strains two distinct enzymes are found to be produced at high levels:

48a936ef-68c5-4d35-81b2-c19ac5469298

1

D-amino acid oxidase (EC 1.4.3.3) and D-tryptophan oxidase. A convenient assay procedure for D-tryptophan oxidase is described. The two enzymes could be distinguished on the basis of their sensitivity to inhibition by L-phenylalanine and L-tyrosine. Strains that were trp dadR could not grow with D-tryptophan in the presence of L-phenylalanine, but further mutations, Fyo, could be isolated that allowed growth under these conditions.

48a936ef-68c5-4d35-81b2-c19ac5469298

2

Some of them were characterized by further increases in the level of D-tryptophan oxidase activity and a sharp decrease in D-amino acid oxidase. These kinds of Fyo mutations lay in or near the dadR gene. The substrate specificity of the two enzymes toward a large number of compounds was examined.

48a936ef-68c5-4d35-81b2-c19ac5469298

3

The transamination of aromatic keto acids was investigated. In the wild-type strain only a single enzyme, transaminase A (EC 2.6.1.5), was found, and it was irreversibly activated when subjected to elevated temperatures. The present state of our knowledge on D-amino acid utilization in E.

48a936ef-68c5-4d35-81b2-c19ac5469298

4

coli is summarized.

74c3405b-1582-4c08-aa62-9679284c4b0c

0

Transduction of chromosomal genes between enteric bacteria by bacteriophage P1. We have used P1 transduction to create intergeneric hybrid strains of enteric bacteria by moving the genA and hut genes between Klebsiella aerogenes, Escherichia coli and Salmonella typhimurium. The use of E. coli as the recipient in such transductions permits the construction of episomes and specialized transducing phage containing non-E. coli material. The effect of host restriction modification and deoxyribonucleic acid homology on the frequency of intergeneric transduction of these loci has been examined.

a9eede7e-25ba-4c7c-a675-4875b88c56a7

0

Galactoside accumulation by Escherichia coli, driven by a pH gradient. Acidification of the external medium results in thiomethylgalactoside accumulation in an energy-depleted adenosine triphosphatase-negative mutant of Escherichia coli.

db033684-25ab-4037-87e2-aa0a6393beef

0

Adenosine 5'-monophosphate-stimulated cyanide-insensitive respiration in mitochondria of Moniliella tomentosa. Mitochondria of the yeastlike fungus Moniliella tomentosa oxidize reduced nicotinamide adenine dinucleotide, reduced nicotinamide adenine dinucleotide phosphate, succinate, isocitrate, and lactate. These oxidations are completely inhibited by cyanide or antimycin A in mitochondria isolated from cells grown in the standard medium.

db033684-25ab-4037-87e2-aa0a6393beef

1

On the other hand, the oxidation of all substrates, except lactate, is almost completely insensitive to cyanide or antimycin A in mitochondria from cells grown in the presence of ethidium bromide. In this instance, the oxidation is mainly mediated by an alternate oxidase which can be blocked by salicyl hydroxamic acid.

db033684-25ab-4037-87e2-aa0a6393beef

2

The alternate oxidase can be specifically stimulated by adenosine 5'-monophosphate and this provides a new method for the characterization of the alternate oxidase in mitochondria of M. tomentosa.

1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e

0

Sulfate-reducing pathway in Escherichia coli involving bound intermediates. Although a sulfate-reducing pathway in Escherichia coli involving free sulfite and sulfide has been suggested, it is shown that, as in Chlorella, a pathway involving bound intermediates is also present. E. coli extracts contained a sulfotransferase that transferred the sulfonyl group from a nucleosidephosphosulfate to an acceptor to form an organic thiosulfate.

1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e

1

This enzyme was specific for adenosine 3'-phosphate 5'-phosphosulfate, did not utilize adenine 5'-phosphosulfate, and transferred to a carrier molecule that was identical with thioredoxin in molecular weight and amino acid composition. In the absence of thioredoxin, only very low levels of the transfer of the sulfo group to thiols was observed.

1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e

2

As in Chlorella, thiosulfonate reductase activity that reduced glutathione-S-SO3- to bound sulfide could be detected. In E. coli, this enzyme used reduced nicotinamide adenine dinucleotide phosphate and Mg2+, but did not require the addition of ferredoxin or ferredoxin nicotinamide adenine dinucleotide phosphate reductase. Although in Chlorella the thiosulfonate reductase appears to be a different enzyme from the sulfite reductase, the E.

1ebff0d0-25f7-4ff0-8cd5-83cf2a51267e

3

coli thiosulfonate reductase and sulfite reductase may be activities of the same enzyme.

dd366adb-2b4f-4d95-a0f5-ba82d756ff7f

0

An endonuclease from Escherichia coli that introduces single polynucleotide chain scissions in ultraviolet-irradiated DNA. An endonuclease that makes single polynucleotide chain scissions in ultraviolet-irradiated DNA has been purified from Escherichia coli. The activity has the following properties: (a) unirradiated DNA is attacked very little if at all;

dd366adb-2b4f-4d95-a0f5-ba82d756ff7f

1

(b) single strand DNA is not attacked, whether irradiated or not; (c) there is no requirement for divalent cations and the activity is not affected by the addition of EDTA; (d) the pH optimum is approximately 7; (e) the activity is inhibited by 1 M NaCl, single strand DNA, transfer RNA and double strand DNA;

dd366adb-2b4f-4d95-a0f5-ba82d756ff7f

2

(f) the sedimentation coefficient, S20,w, is approximately 2.6; (g) it is a basic protein. The enzyme is tentatively named E. coli endonuclease III. The physiological function of the endonuclease has not yet been established.

4269d979-ab90-4056-b518-4eced3f4b7dd

0

Quaternary conformational changes in human hemoglobin studied by laser photolysis of carboxyhemoglobin. These experiments indicate that absorbance changes observed at the 425 nm isosbestic point of the Hb and HbCO following laser photolysis of HbCO provide a direct measure of the rates of quaternary conformational changes between rapidly reacting Hb (the immediate product of full photolysis) and slowly reacting normal deoxyhemoglobin.

4269d979-ab90-4056-b518-4eced3f4b7dd

1

Hb, first observed by Gibson (Gibson, Q.H. (1959) Biochem. J. 71, 293-303), Has been interpreted as deoxyhemoglobin remaining in the liganded quaternary conformation following rapid removal of ligand by a light pulse. In borate buffers between pH 8.4 and 9.6 particularly simple pH-independent results were obtained which allowed the use of a Monod.

4269d979-ab90-4056-b518-4eced3f4b7dd

2

Wyman, and Changeux model (Monod, J., Wyman, J., and Changeux, J (1965) J. Mol. Biol. 12, 88-118) to fit the data. In this case Hb is taken to be R state deoxyhemoglobin. Partial photolysis experiments at 425 nm show that the rate of the R - T conformational change at 20 degrees decreases by about a factor of 2 for each additional bound ligand.

4269d979-ab90-4056-b518-4eced3f4b7dd

3

The rate of the ligand-free conformational change is found to be 920 +/- 60s(-1), 6400 +/- 600s(-1), and 15,700 +/- 700(-1) respectively at 3 degrees, 20 degrees, and 30 degrees. The previously uninterpreted effects of flash length and partial photolysis on the CO recombination kinetics can be explained in terms of the present model.

4269d979-ab90-4056-b518-4eced3f4b7dd

4

Kinetic results obtained below pH 8 are found to be inconsistent with a two-state model. It appears that binding of inositol hexaphosphate produces a new rapidly reacting quaternary conformation of HbCO.

ecfd0bd1-87f5-447c-ae0d-1e2815db0c9b

0

Muscarinic acetylcholine receptor from rat brain. Partial purification and characterization. A protein capable of binding atropine and (3H)propylbenzilylcholine mustard was solubilized and purified (200-fold) from rat brain. Pronase and trypsin, but not phospholipases, diminished the binding capacity of the solubilized receptor. The molecular weight of the salt-solubilized receptor as determined by gel filtration in the absence of detergents is 30,000.

ecfd0bd1-87f5-447c-ae0d-1e2815db0c9b

1

The purified protein showed specificity of binding toward muscarinic ligands. the high and low affinity dissociation constants of the receptor.atropine complex are 0.3 nM and 0.15 muM. Binding of atropine is pH-dependent with an optimum at 7.1. Ca2+ influences the binding of atropine and maximal binding occurs at 0.5

ecfd0bd1-87f5-447c-ae0d-1e2815db0c9b

2

mM Ca2+. The subcellular distribution of the receptor was also examined.

5e00ac47-ef47-4711-9088-6c6887b5b416

0

Quantitative relationships between phosphorylation, electron flow, and internal hydrogen ion concentrations in spinach chloroplasts. 1. Further evidence that the uptake of [14C]hexylamine, determined by centrifugal filtration of spinach chloroplast thylakoids through silicone fluid layers, gives precise estimations of light-induced H+ concentration gradients (deltapH) is presented.

5e00ac47-ef47-4711-9088-6c6887b5b416

1

DeltapH was independent of the amount of thylakoids used or of the concentration of hexylamine. Moreover, hexylamine uptake was sensitive to the osmolarity of the suspending medium. 2. Internal H+ concentration ([H+]in) is proportional to the rate of electron flow when light intensity was used to vary these parameters.

5e00ac47-ef47-4711-9088-6c6887b5b416

2

Proportionality was still observed in the presence of 0.1 and 1.0 muM gramicidin D. When, however, [H+]in and electron flow were varied by increasing the concentration of gramicidin D, at constant light intensity the rate of electron flow was approximately proportional to 1/[H]in.

5e00ac47-ef47-4711-9088-6c6887b5b416

3

3. The phosphorylation efficiency (P/e2 ratio) falls with decreasing light intensity or increasing concentrations of the phosphorylation inhibitor, 4'-deoxyphlorizin. The proportionality between the rate of electron flow and [H+]in allows the calculation of the rate of nonphosphorylating (basal) electron flow if [H+]in under phosphorylating conditions is known.

5e00ac47-ef47-4711-9088-6c6887b5b416

4

The contribution of basal electron flow, a consequence of passive efflux of H+ from the thylakoids, to the overall rate of electron flow increases as the rate of phosphorylation decreases. P/e2 ratios calculated using rates of electron flow from which the basal component has been subtracted are constant.

5e00ac47-ef47-4711-9088-6c6887b5b416

5

A calculated P/e2 ratio of about 1.3 is obtained. 4. It is shown that the reciprocal of the phosphorylation efficiency should be proportional to 1/[H+]in2 when these parameters are varied using light intensity. This relationship was verified and provided an estimate of the P/e2 at infinite [H+]in.

5e00ac47-ef47-4711-9088-6c6887b5b416

6

This value was 1.3. These results provide further evidence that a H+ electrochemical gradient serves to couple photophosphorylation to electron flow and that the rate of phosphorylation is proportional to [H+]in3. That is, three H+ are translocated out of thylakoids for each adenosine triphosphate formed.

c34b4ecf-680c-497c-9ff0-fad105f6120b

0

Kinetic properties of human placental aromatase. Application of an assay measuring 3H2O release from 1beta,2beta-3H-androgens. The rapid and sensitive assay of 1beta,2beta-3H-androgen aromatization by measurement of 3H2O release (Thompson, E.A., Jr., and Siiteri, P.K. (1974) J. Biol.

c34b4ecf-680c-497c-9ff0-fad105f6120b

1

Chem. 249, 5364-5372) has been analyzed to determine its applicability to initial rate studies. It was found that aromatization is the sole reaction catalyzed by lyophilized placental microsomes that causes a loss of tritium from position 1 or 2 of androstenedione and testosterone. Tritium is, however, removed from position 2 of the estrogen products, presumably in 2-hydroxylation, but this does not invalidate use of the assay for initial rate measurements;

c34b4ecf-680c-497c-9ff0-fad105f6120b

2

it was therefore used to characterize the catalytic properties of aromatase. Aromatization by the freeze-dried preparation was stimulated by K+, EDTA, and dithiothreitol, and was maximally active at pH 7.5 TO 8.0. With incubation conditions optimized for these factors, the apparent Km for NADPH is approximately 1 muM.

c34b4ecf-680c-497c-9ff0-fad105f6120b

3

The maximum velocity of androstenedione aromatization exceeds that of testosterone, and the affinity of the substrate binding site is higher for the former substrate, the apparent Km values being 0.1 muM and 0.4 muM, respectively. Mutual competition experiments with the androgen substrates showed that each gives simple competitive inhibition of the other's aromatization;

c34b4ecf-680c-497c-9ff0-fad105f6120b

4

furthermore, the apparent Ki values for each are in close agreement with their respective Km values. Androst-1,4,6-triene-3,17-dione competitively inhibits the aromatization of both androstenedione and testosterone, the apparent Ki, in both cases being 0.2 muM. It is concluded that the two androgen substrates are aromatized at a single, identical site.

9471d7ca-281e-466f-957f-5f8ef72b4aca

0

Apparent oxidation-reduction potential of Clostridium acidi-urici ferredoxin. Effect of pH, ionic strength, and amino acid replacements. The effects of pH and ionic strength on the midpoint reduction potential (Emp) of Clostridium acidi-urici ferredoxin were determined using hydrogen gas and hydrogenase. The Emp of native ferredoxin at 24-25 degrees in 0.1

9471d7ca-281e-466f-957f-5f8ef72b4aca

1

M Tris-chloride buffer, pH 7.0, is--0.434 V. In the pH range examined, the Emp becomes approximately 13 mv more negative per each pH unit increase. A plot of the log of ionic strength versus the apparent Emp of ferredoxin in 0.1 M Tris-chloride buffer, pH 7.5

9471d7ca-281e-466f-957f-5f8ef72b4aca

2

, Was linear over the range of 1.0 to 0.01 ionic strength with Emp values of--0.414 and--0.475 V, respectively, at these extremes. This effect is the same with sodium chloride, sodium bromide, or ammonium sulfate. Potassium phosphate buffer caused a similar change, but the absolute values of Emp differed from those obtained in the presence of the other salts.

9471d7ca-281e-466f-957f-5f8ef72b4aca

3

This effect of pH and ionic strength on Emp may be general for clostridial-type (Fe4S4)2-ferredoxins, since the apparent Emp of Clostridium pasteurianum ferredoxin is affected in a similar manner by these two variables. The Emp of this ferredoxin in 0.1 M Tris-chloride buffer pH 7.0

9471d7ca-281e-466f-957f-5f8ef72b4aca

4

, is--0.405 V. Since the NH2-terminal amino acid residue, Ala1, and Tyr2 of C. acidi urici ferredoxin are near an (Fe4S4)2-cluster in the protein, the apparent Emp of derivatives that contained amino acid replacements in these two positions were determined. Under similar conditions, the Emp of most of the 13 derivatives examined, including those of [Leu2]- and[3-NH2-Tyr30]ferredoxin, is approximately the same as that of native ferredoxin.

9471d7ca-281e-466f-957f-5f8ef72b4aca

5

However, the Emp of [His2]ferredoxin is approximately 15 mv more positive, whereas that of [Trp2]ferredoxin is 22 mv more negative than that of native C. acidi-urici ferredoxin. Variations in sodium chloride concentration and pH also affected the apparent Emp of the derivatives. It is suggested that the changes observed in the Emp of C.

9471d7ca-281e-466f-957f-5f8ef72b4aca

6

acidi-urici ferredoxin are caused by protein conformational changes.

c42335f1-02b8-471d-ad22-742cf535d1ba

0

A deoxyribonucleic acid kinase from nuclei of rat liver. Purification and properties. A DNA kinase has been partially purified from rat liver nuclei by a procedure which also yields DNA ligase. The kinase uses ATP to phosphorylate specifically the 5'-hydroxyl termini of oligodeoxynucleotides and of single- or double-stranded DNA, yielding 5'-phosphate termini and ADP.

c42335f1-02b8-471d-ad22-742cf535d1ba

1

The kinase is inactive on RNA, or on oligodeoxynucleotides of chain length less than approximately 10 to 12 residues. The kinase requires a divalent cation (Mg2+, Mn2+, Co2+, Zn2+, Ni2+, or Ca2+) for activity and has an acidic pH optimum. It is inhibited by a variety of nucleotides as well as by very low levels of inorganic and organic sulfate compounds and sulfate analogues.

c42335f1-02b8-471d-ad22-742cf535d1ba

2

The molecular weight of the kinase is estimated to be 8 times 10(4) from gel filtration.

e7c1efed-2035-48a9-b97f-bd8cdd33e243

0

Hydrogen exchange at the amide group of reduced pyridine nucleotides and the inhibition of that reaction by dehydrogenases. Stopped flow ultraviolet spectroscopy has been used to measure the rate of hydrogen exchange with solvent at the amide group of reduced nicotinamide nucleotide coenzymes. Several mechanisms for the exchange reaction are considered in the light of the kinetic data. Complex formation between the coenzyme and any of four dehydrogenases markedly slows the rate of hydrogen exchange. Hydrogen bond formation and/or hydrophobic interactions within these complexes are thought to be the reasons for the decreased rate of exchange.

f2437d01-97be-470f-afc0-2ca92f82b5ff

0

A Dacron wool packed-bed extracorporeal reactor: a kinetic study of immobilized Escherichia coli II L-asparaginase. An extracorporeal reactor containing a packed bed of Dacron fibers has been developed. Escherichia coli II L-asparaginase was coupled to the Dacron using gamma-aminopropyltriethoxysilane and glutaraldehyde. The preparation had an activity of 37 IU per gram of Dacron (37 degrees C).

f2437d01-97be-470f-afc0-2ca92f82b5ff

1

The apparent Km was studied as a function of the flow rate. The data indicated that the apparent Km approached the Km of the native enzyme at flow rates of about 300 mg/min. In vivo use of L-asparaginase immobilized on the Dacron indicated effective lowering of plasmatic L-asparagine levels.

653d2110-ff66-465a-b3ec-a0940565fa06

0

Isolation of intact megakaryocytes from guinea pig femoral marrow. Successful harvest made possible with inhibitions of platelet aggregation; enrichment achieved with a two-step separation technique. Methods have been devised to harvest megakaryocytes from guinea pig femoral marrow and to isolate them in high yield. When marrow tissue was disaggregated the megakaryocytes underwent degenerative changes characterized by the loss of cytoplasmic granules and alterations in membrane topography, similar to the changes seen in aggregating platelets.

653d2110-ff66-465a-b3ec-a0940565fa06

1

These morphologic changes were interpreted to mean that megakaryocytes possessed functional attributes of platelets. The use of agents which inhibit platelt aggregation (0.38% sodium citrate. 10(-3) M adenosine, and 2 x 10(-3) M theophylline) in a medium free of bivalent cations prevented these changes.

653d2110-ff66-465a-b3ec-a0940565fa06

2

This solution resulted in both an excellent morphologic preservation and a significantly increased recovery of megakaryocytes from marrow tissue. A two-step purification of the intact megakaryocytes was carried out on the basis of their low density and large size, with equilibrium density gradient centrifugation followed by velocity sedimentation.

653d2110-ff66-465a-b3ec-a0940565fa06

3

This sequence gave approximately a 100-fold enrichment of megakaryocytes, significantly better than that achieved with either method alone. These techniques for harvesting and concentrating megakaryocytes make it possible for the first time to study megakaryocytes in vitro.

b619ca86-18fe-4f4d-85c9-1250584d54e1

0

The polymerization of actin. III. Aggregates of nonfilamentous actin and its associated proteins: a storage form of actin. When echinoderm sperm are treated with the detergent Triton X-100 at pH 6.4 in 10 mM phosphate buffer, the membranes are solubilized, but the actin which is located in the periacrosomal region remains as a phase-dense cup.

b619ca86-18fe-4f4d-85c9-1250584d54e1

1

These cups can be isolated free from the flagella and chromatin and can be solubilized by increasing the pH to 8.0 and by changing the ionic strength and type of buffer used. Since the actin does not exist in the "F" state in unreacted sperm, and since the actin remains as a unit that does not diffuse away, it must be present in the mature sperm in a bound or storage state.

b619ca86-18fe-4f4d-85c9-1250584d54e1

2

The actin is, in fact, associated with a pair of proteins whose mol wt are 250,000 and 230,000. When the isolated cups are digested with trypsin, these high molecular weight proteins are digested, thereby liberating the actin. The actin will polymerize if heavy meromyosin or subfragment 1 is added to a preparation of isolated cups.

b619ca86-18fe-4f4d-85c9-1250584d54e1

3

Evidence is presented that this pair of high molecular weight proteins is similar in molecular weight and properties to erythrocyte spectrin. Attempts at transforming the storage form of actin in the cup into filaments were only moderately successful. The best conditions for filament formation involve incubating the cup in ATP and divalent salts.

b619ca86-18fe-4f4d-85c9-1250584d54e1

4

Careful examination of these cups reveals that the actin polymerized preferentially on either end of oriented filaments that already exist in the cup, indicating that self-nucleation is inefficacious. I conclude that the actin can exist in the storage form by its association with spectrin-like molecules and that the actin in this state polymerizes preferentially onto existing filaments.

8c62887e-22a7-4bad-8764-aa1f00cedba6

0

Degradation of abnormal proteins in HeLa cells. The experiments show that abnoramal proteins are degraded faster than normal ones in HeLa cells. Among the fragmentary proteins made in the presence of puromycin, those with low molecular weight are least stable. Proteins made after incubation with 5-fluorouracil or in the presence of some amino acid analogues are also unstable. Breakdown of proteins made in the presence or absence of puromycin is nearly unaffected by cycloheximide and is independent of pH between 7 and 8.

44905afb-5ef8-48c7-8d10-5a09806bf399

0

The effect of pH on incorporation of galactose by a normal human cell line and cell lines from patients with defective galactose metabolism. Incorporation of radioactive galactose into TCA-insoluble material of galactosemic fibroblasts is more sensitive to low pH than is the incorporation by normal human fibroblasts.

44905afb-5ef8-48c7-8d10-5a09806bf399

1

This study was undertaken to determine (1) whether there was any pH which could correct or counteract the galactosemic defect relative to galactose incorporation, and (2) whether the low pH effect was specific for galactose metabolism or whether general cellular metabolism in galactosemic cells was more sensitive to low pH than that in normal cells. The pH dependencies of incorporation of radioactive galactose and glucose into cellular macromolecules were investigated in galactosemic and normal cells.

44905afb-5ef8-48c7-8d10-5a09806bf399

2

Normal cells have a biphasic curve with respect to galactose incorporation with peaks at pH 7.0 and 8.5. Galactosemic cells have only the high pH peak. The maximum incorporation by galactosemic cells was never more than about 30% that seen by normal cells under the conditions of these experiments.

44905afb-5ef8-48c7-8d10-5a09806bf399

3

Thus manipulation of the pH alone cannot correct the galactosemic defect. The rate of incorporation of radioactive galactose was studied in normal, galactosemic and galactokinase deficient cells, at pH 7.2 and at pH 6.3. At pH 7.2, galactosemic cells incorporate galactose at a linear rate which is 30 to 40% that of normal cells while incorporation by kinase-deficient cells is between 5 and 10% of normal.

44905afb-5ef8-48c7-8d10-5a09806bf399

4

At pH 6.3, the incorporation is also linear. However, galactosemic cells now exhibit the same rate as kinase-deficient cells in which the low level of incorporation is unaffected by pH. These results suggest that incorporation of galactose by galactosemic cells at low pH is not due to metabolic death of the cells, but may be due to the inhibition of some specific step or steps along a metabolic route of galactose metabolism other than the Leloir pathway.

ce1b057b-8304-41d7-9ae3-869260b84d02

0

Chromatography of hemoglobins on CM-cellulose with bis-tris and sodium chloride developers. CM-Cellulose as an ion-exchange medium with Bis-tris as buffer and a gradient of sodium chloride provides a versatile system for the chromatography of hemoglobins. Changes in pH, Bis-tris concentration, and slope of the sodium chloride grandient provide means for markedly altering chromatographic behavior for special separations. Examples are given of the application of the method to normal samples and to those with hemoglobinopathies.

110fcb13-2778-42f6-9e92-d0b32261ae35

0

The determination of phanquone in biological material by gas-liquid chromatography. A gas-liquid chromatographic method for the quantitative determination of phanquone is described, based on the formation of a dimethoxine prior to its extraction from biological material. The sensitivity of the procedure is about 15 ng/ml in biological fluid.

299ae760-af48-4113-91f6-35053fe45eb6

0

Stability of fluorescent antibody conjugates stored under various conditions. Two experiments were carried out to determine the stability of fluorescent antibody conjugates. In experiment 1, Francisella tularemia conjugates in the lyophilized state retained their original staining titer for 1,294 days when stored at 25, 4 to 5, and -20 C;

299ae760-af48-4113-91f6-35053fe45eb6

1

at 37 C the conjugates were stable for at least 65 days. In the liquid state at pH 7.4 and 8.0 these conjugates were stable for 1,294 days at 4 to 5 and -20 C, whereas those stored at 25 C remained stable through days 473 and 160 of storage, respectively, after which the staining titer gradually dropped.

299ae760-af48-4113-91f6-35053fe45eb6

2

In experiment 2 five previously lyophilized conjugates were rehydrated with three different diluents and stored at 4 to 5 C for up to 600 days at their working dilutions. All of these conjugates retained their original staining titer during the test period except an anti-human globulin conjugate rehydrated with phosphate-buffered saline.

299ae760-af48-4113-91f6-35053fe45eb6

3

Recommendations are made for the long-term storage of fluorescent antibody conjugates.

22dcea31-f492-4856-bcd1-c87458fcc1d0

0

Inactivation of viruses in serum with binary ethyleneimine. The inactivation os six strains from three different groups of viruses with 0.001 M binary ethyleneimine at 37 C proceeded at the same rate in either bovine serum or cell culture medium. The inactivant did not impair the growth-promoting capacity of bovine serum used in cell culture, nor did it affect the antibody activity of guinea pig hyperimmune serum.

c457bed7-c03a-4af8-89bf-d92bbdca4d44

0

Inorganic phosphate homeostasis. Renal adaptation to the dietary intake in intact and thyroparathyroidectomized rats. The possibility of renal tubular adaptation to variations in dietary inorganic phosphate (Pi) was investigated in intact and thyroparathyroidectomized (TPTX) rats pair-fed diets containing low, normal, and high amounts of Pi for periods up to 10 days.

c457bed7-c03a-4af8-89bf-d92bbdca4d44

1

Clearances were measured before and during active i.v. infusions with Pi in conscious animals. Thus tubular reabsorption of phosphate (TRPi) could be assessed over a wide range of plasma phosphate concentrations ([Pi]P1). It was found that the renal tubule could adapt its capacity to transport Pi according to the dietary Pi:

c457bed7-c03a-4af8-89bf-d92bbdca4d44

2

TRPi was always higher, for a given [Pi]P1, in the animals fed low than in those fed higher Pi diets. This diet-induced modification also occurred in the absence of thyroparathyroid glands, in the presence of the same calcemia and urinary pH, and during marked extracellular volume expansion.

c457bed7-c03a-4af8-89bf-d92bbdca4d44

3

A time-course study in rats TPTX both before and during the administration of the experimental diets showed that a difference in the tubular handling of Pi was detectable as early as 3 days after switching the animals from a normal to low- or high-Pi diets. These results indicate that factors other than parathyroid hormone are implicated in the tubular response to variations in the dietary intake of inorganic phosphate.

247316a1-e4e7-4395-8a87-e3ef37de5923

0

Effect of beta adrenergic blockade on renin response to renal nerve stimulation. The ability of d,l-propranolol to block renin secretion in response to various extrarenal stimuli, such as hemorrhage and hypoglycemia, has been interpreted to indicate the presence of an intrarenal beta receptor regulating renin release. However, two problems complicate this interpretation:

247316a1-e4e7-4395-8a87-e3ef37de5923

1

(a) the stimuli have effects outside the kidney, and (b) d,l-propranolol has a local anesthetic, as well as a beta adrenergic blocking, action. In the present study, the effects of a purely intrarenal stimulus, in the form of renal nerve stimulation (RNS), on renin secretion was examined.

247316a1-e4e7-4395-8a87-e3ef37de5923

2

The effects of d,l-propranolol (anesthetic and beta-blocking activity), l-propranolol (beta-blocking activity only), and d-propranolol (local anesthetic activity only) on the renin response to RNS were examined. In a control group of animals, two sequential RNS increased mean renin secretion from 401 to 1,255 U/min (P less than 0.25

247316a1-e4e7-4395-8a87-e3ef37de5923

3

) and from 220 to 2,179 U/min (P less than 0.01). In a second group the first RNS increased renin secretion from 201 to 1,181 U/min (P less than 0.01), but after d,l-propranolol was given RNS did not significantly alter renin secretion (33 to 55 U/min).

247316a1-e4e7-4395-8a87-e3ef37de5923

4

In a third group the initial RNS increased renin secretion from 378 to 1,802 U/min (P less than 0.025), but after l-propranolol was given RNS had no significant effect on renin secretion (84 to 51 U/min). A fourth group of dogs showed a rise in renin secretion from 205 to 880 U/min (P less than 0.001

247316a1-e4e7-4395-8a87-e3ef37de5923

5

) in response to the first RNS, while the second RNS, given after an infusion of d-propranolol, caused a rise in renin secretion from 80 to 482 (P less than 0.005). The nature of the electrical stimulus was consistent in all groups and caused no detectable changes in renal or systemic hemodynamics or in urinary electrolyte excretion.

247316a1-e4e7-4395-8a87-e3ef37de5923

6

The results, therefore, indicate that renin secretion can be stimulated through intrarenal beta receptors independent of changes in systemic or renal hemodynamics or in tubular sodium reabsorption. Hence the effect of beta stimulation on renin secretion would appear to result from a direct action on the renin-secreting cells of the juxtaglomerular apparatus.