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0145255f-035e-4060-94a2-944d744fcfef
| 1 |
The effects of reaction with 2-hydroxy-5-nitrobenzyl bromide, tetranitromethane and hydrogen peroxide have been studied. Conversely, treatment of immobilized antibodies with 2-hydroxy-5-nitrobenzyl bromide facilitates the elution of glucagon during immunoaffinity chromatography. The general implications of these results are discussed.
|
ac39aee5-dd99-4935-9590-fe71ab4c2e33
| 0 |
Release and activation of a particulate bound acid phosphatase from Tetrahymena pyriformis. A sedimentable form of acid phosphatase (EC 3.1.3.2) from Tetrahymena pyriformis was found to be solubilized by Triton X-100. The total enzyme activity in the insoluble cell fraction increased almost 200% upon solubilization with Triton X-100 or Nonidet P-40.
|
ac39aee5-dd99-4935-9590-fe71ab4c2e33
| 1 |
Removal of membrane lipids and Triton X-100 from the particulate wash solution with a chloroform extraction resulted in non-specific enzyme-protein aggregation which was reversible upon addition of Triton X-100. The results indicate that this acid phosphatase is an integral membrane protein. The pH optima for this particulate bound acid phosphatase was 3.5
|
ac39aee5-dd99-4935-9590-fe71ab4c2e33
| 2 |
with o-carboxyphenyl phosphate and 4.0 with p-nitrophenyl phosphate as substrates. The Km values of each substrate were 3.1 and 0.031 mM, respectively.
|
73852e01-16a9-48b4-8bab-4ddc180f5773
| 0 |
Partial purification and characterization of post-proline cleaving enzyme: enzymatic inactivation of neurohypophyseal hormones by kidney preparations of various species. The inactivation of the neurohypophyseal hormones arginine vasopressin and oxytocin, both 14C-labelled in the C-terminal glycine residue, by enzymes present in kidney homogenates of various species has been investigated, and some of the enzymes responsible have been partially purified and characterized.
|
73852e01-16a9-48b4-8bab-4ddc180f5773
| 1 |
The Leu-Gly peptide bond of oxytocin is generally most effectively cleaved by kidney homogenates, although with certain species enzymic activity hydrolyzing the Pro-Leu bond is significant. Degradation of arginine vasopressin is slower than oxytocin in all species studied, and appears to occur by a different overall mechanism since cleavage of the Pro-Arg bond is more significant than hydrolysis of the Arg-Gly bond.
|
73852e01-16a9-48b4-8bab-4ddc180f5773
| 2 |
The enzyme releasing glycinamide from oxytocin and the "Post-Proline Cleaving Enzyme", which releases C-terminal dipeptide from oxytocin and arginine vasopressin, were partially purified from lamb kidney by ammonium sulfate fractionation and column chromatography. The two enzymes are shown to be separate entities with different pH profiles.
|
73852e01-16a9-48b4-8bab-4ddc180f5773
| 3 |
The prolyl peptidase activity released the C-terminal dipeptides from oxytocin and arginine vasopressin at similar rates and was inhibited by p-chloromercuriphenylsulfonic acid, 1,10-phenanthroline, L-1-tosylamido-2-phenylethylchloromethyl ketone, Co2+, Ca2+, and Zn2+, but significantly enhanced by dithiothreitol. The prolyl peptidase preparation cleaves proline-containing peptide substrates at the Pro-X bond.
|
73852e01-16a9-48b4-8bab-4ddc180f5773
| 4 |
The rate of cleavage is dependent on the nature of residue X and with the conditions used there is no cleavage when X equals Pro; however, cleavage occurs when X is a D isomer: [Mpr1, D-Arg8] vasopressin is inactivated at a rate similar to [Mpr1, Arg8]- and [Mpr1, Lys8] vasopressin, suggesting that the known prolonged biological action of [Mpr1, D-Arg8] vasopressin is not due to resistance to the prolyl peptidase.
|
73852e01-16a9-48b4-8bab-4ddc180f5773
| 5 |
In all characteristics tested the lamb kidney prolyl peptidase was identical to the post-proline cleaving enzyme isolated earlier from human uterus. In vivo experiments in the cat suggested that both the glycinamide-releasing enzyme and post-proline cleaving enzyme are present and effective in inactivating neurohypophyseal hormones in the intact animal.
|
02ac7e6c-fcfa-4019-bd3c-b7037eddf187
| 0 |
Adenosine phosphyorylase activity as distinct from inosine-guanosine phosphorylase activity in Sarcoma 180 cells and rat liver. Adenosine phosphorylase (EC 2.4.2.-) activity present in Sarcoma 180 cells grown in culture and in rat liver, is shown to be distinct from inosine-guanosine phosphorylase by several criteria:
|
02ac7e6c-fcfa-4019-bd3c-b7037eddf187
| 1 |
(a) treatment of Sarcoma 180 cell extract with p-chloromercuribenzoate inhibited the two activities to a different extent, (b) adenine selectively protected the adenosine phosphorylase activity of Sarcoma 180 and rat liver extract against heat inactivation, while hypoxanthine selectively protected inosine-guanosine phosphorylase activity, (c) at nearly saturating substrate concentrations and using Sarcoma 180 extract, the rates of ribosylation of a mixture of adenine + hypoxanthine or adenine + guanine, but not of hypoxanthine + guanine, were found to be almost equal to the sum of their individual rates as measured separately, (d) inosine selectively inhibited the ribosylation of hypoxanthine and guanine catalysed by Sarcoma 180 and rat liver extract while 2-chloroadenosine selectively inhibited the ribosylation of adenine and N6-furfuryladenine, (e) pH vs.
|
02ac7e6c-fcfa-4019-bd3c-b7037eddf187
| 2 |
activity curves were similar with hypoxanthine or guanine as the substrate but they were markedly different from the curve with adenine as the substrate. The potential role of adenosine phosphorylase activity in vivo is discussed.
|
8624b51e-85c0-4f29-8b9a-ac4094628570
| 0 |
Purification and properties of an alpha-amylase inhibitor from wheat. Four inhibitors of alpha-amylase (EC 3.2.1.1) were separated from an alcohol extract of wheat by ion-exchange chromatography on DE52-cellulose. One inhibitor, which showed the greatest specificity for human salivary amylase relative to human pancreatic amylase, has been purified by the following steps:
|
8624b51e-85c0-4f29-8b9a-ac4094628570
| 1 |
(a) alcohol fractionation (60--90%) of water extract (b) ion-exchange chromatography on QAE-Sephadex A-50; (c) re-chromatography on DE52-cellulose and (d) gel filtration on Sephadex G-50. The purified inhibitor is 100 times more specific for human salivary amylase than for human pancreatic amylase.
|
8624b51e-85c0-4f29-8b9a-ac4094628570
| 2 |
It shows an electrophoretic mobility of 0.2 on disc gel electrophoresis and a molecular weight of about 21 000. This inhibitor contributes about 16% to the total salivary amylase inhibiting power of the wheat extract.
|
512361cc-51ac-4e26-a17a-481f1c4f2138
| 0 |
Purification and specificity of prolyl dipeptidase from bovine kidney. Prolyl dipeptidase (iminodipeptidase, L-prolyl-amino acid hydrolase, EC 3.4.13.8) was purified 180-fold from bovine kidney. The enzyme which was obtained in a 10% yield was completely separated from a number of known kidney peptidases including an enzyme of very similar substrate specificity, proline aminopeptidase (L-prolyl-peptide hydrolase, EC 3.4.11.5
|
512361cc-51ac-4e26-a17a-481f1c4f2138
| 1 |
). The specific activity of the enzyme with L-prolylglycine as substrate is 1600 units of activity per mg protein. Optimum activity of the enzyme is at pH 8.75 and the molecular weight on gel filtration was estimated to be 100 000. The isoelectric point of the enzyme is pH 4.25
|
512361cc-51ac-4e26-a17a-481f1c4f2138
| 2 |
. Studies of substrate specificity showed that the enzyme preferentially hydrolyzes dipeptides and dipeptidyl amides with L-proline or hydroxy-L-proline at the N-terminus. Longer chain substrates with N-terminal proline were not hydrolyzed.
|
8a52fcbe-111f-4d65-971b-da9ce2554ea9
| 0 |
Isolation and characterization of beta-glucosidase from the cytosol of rat kidney cortex. A procedure is described for the preparation of extensively purified beta-D-glucosidase (EC 3.2.1.21) from the cytosol fraction of rat kidney. The specific activity of the beta-glucosidase in the high speed supernatant (100 000 X g, 90 min) fraction of rat kidney homogenate is 700-fold greater than that in the same fraction from heart, skeletal muscle, lung, spleen, brain or liver.
|
8a52fcbe-111f-4d65-971b-da9ce2554ea9
| 1 |
beta-Glucosidase activity co-chromatographs with beta-D-galactosidase, beta-D-fucosidase, alpha-L-arabinosidase and beta-D-xylosidase activities through the last four column steps of the purification and their specific activities are 0.26, 0.39, 0.028 and 0.017 relative to that of beta-glucosidase, respectively. The specific activity of the apparently homogeneous beta-glucosidase is 115 000 nmol of glucose released from 4-methylumbelliferyl-beta-D-glucopyranoside per mg protein per h.
|
8a52fcbe-111f-4d65-971b-da9ce2554ea9
| 2 |
All five glycosidase activities possess similar pH dependency (pH optimum, 6--7) and heat lability, and co-migrate on polyacrylamide disc gels at pH 8.9 (RF, 0.67). beta-Glucosidase acitivity is inhibited competitively by glucono-(1 leads to 5)-lactone (KI, 0.61 mM) and non-competitively by a variety of sulfhydryl reagents including N-ethylmaleimide, p-chloromercuribenzoate, 5,5'-dithio-bis(2-nitrobenzoic acid), and iodoacetic acid.
|
8a52fcbe-111f-4d65-971b-da9ce2554ea9
| 3 |
Although the enzyme will release glucose from p-nitrophenyl and 4-methylumbelliferyl derivatives of beta-D-glucose, it will not hydrolyze xylosyl-O-serine, beta-D-glucocerebroside, lactose, galactosylovalbumin or trehalose. The enzyme consists of a single polypeptide chain with a molecular weight of 50 000--58 000, has a sedimentation coefficient of 4.41
|
8a52fcbe-111f-4d65-971b-da9ce2554ea9
| 4 |
S and contains a relatively large number of acidic amino acids. A study of the distribution of beta-glucosidase activity in various regions of the dissected rat kidney indicates that the enzyme is probably contained in cells of the proximal convoluted tubule. The enzyme is also present in relatively large amounts in the villus cells, but not crypt cells, of the intestine.
|
8a52fcbe-111f-4d65-971b-da9ce2554ea9
| 5 |
The physiological substrate and function of the enzyme are unknown.
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 0 |
Studies on (Na+ + K+)-activated ATPase. XXXVIII. A 100 000 molecular weight protein as the low-energy phosphorylated intermediate of the enzyme. Phosphorylation of NaI-treated bovine brain cortex microsomes by inorganic phosphate in the presence of Mg2+ and ouabain has been studied at 0 degrees C (pH 7.4
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 1 |
) and 20 degrees C (pH 7.0). Nearly maximal (90%) and half-maximal phosphorylation are achieved at 20 degrees C within 2 min with 50--155 and 5.6--17 muM 32Pi, respectively, and at 0 degrees C within 75 s with 300--600 and 33--66 muM 32Pi, respectively.
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 2 |
Maximal phosphorylation yields 146 pmol 32P - mg-1 protein. Without ouabain (20 degrees C, pH 7.0) less than 25% of the incorporation observed in the presence of ouabain is reached. Preincubation of the native microsomes with Mg2+ and K+, in order to decompose possibly present high-energy phosphoryl-bonds prior to ouabain treatment, does not affect the maximal phosphate incorporation.
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 3 |
This indicates that the inorganic phosphate incorporation is not due to an exchange with high-energy phosphoryl-bonds, which might have been preserved in the microsomal preparations. Phosphorylation of the native microsomes by ATP in the presence of Mg2+ and Na+ reaches 90 and 50% maximal levels within 15--30 s at 0 degrees C and pH 7.4
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 4 |
at concentrations of [gamma-32P]ATP of 5--32 and 0.5--3.5 muM, respectively. The maximal phosphorylation level is 149 pmol 32P-mg-1 protein, equal to that of ouabain-treated microsomes phosphorylated by inorganic phosphate. Both inorganic phosphate and ATP phosphorylate on site per active enzyme subunit of 135 000 molecular weight.
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 5 |
From the equilibrium constants for the phosphorylation of ouabain-treated microsomes by inorganic phosphate at 0 degrees C and 20 degrees C standard free-energy changes of --5.4 and --6.8 kcal/mol, respectively, are calculated. These values yield a standard enthalpy change of 14 kcal/mol and an entropy change of 70 cal/mol - degree K.
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 6 |
This characterizes the reaction as a process driven by an entropy change. The intermediate formed by phosphorylation with Pi has maximal stability at acidic pH, as is the case for the intermediate formed with ATP. Solubilization in sodium dodecyl sulfate stabilizes the phosphoryl-bond in the pH range of 4--7.
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 7 |
The non-solubilized preparation has optimal stability at pH 2--4, the level of which is equal to that of detergent-solubilized intermediate. Sodium dodecyl sulfate gel electrophoresis of the microsomes at pH 3, following incorporation of 32Pi yields 11 protein bands, only one of which (mol.
|
cdc37f15-5aa1-44b2-b6e3-c28eb2ea5a56
| 8 |
wt 100 000--106 000) carries the radioactive label. This protein has the same molecular weight as the protein, which is phosphorylated by ATP in the presence of Mg2+ and Na+.
|
5e306f9b-61e0-437e-a656-412680ba981f
| 0 |
Partial purification and properties of a chromatin-associated phosphoprotein kinase from rat liver nuclei. A phosphoprotein kinase (EC 2.7.1.37) KIVb, from rat liver nuclei, was purified 75-fold by phosphocellulose chromatography and gel filtration on Sephadex G-200. The enzyme, which has an apparent molecular weight of 55 000, phosphorylates casein and chromatin-bound nonhistone proteins more readily than histones or ribosomal proteins.
|
5e306f9b-61e0-437e-a656-412680ba981f
| 1 |
It exhibits an absolute requirement for divalent cation with optimum activity at 15--20 mM Mg2+. Maximal kinase activity is achieved at 100 mM NaCl. The pH vs. activity curve is biphasic with optima at pH 6.5 and pH 8.0. The Km value for casein is 280 mug/ml and the Km for ATP is 6-10(-6) M.
|
5e306f9b-61e0-437e-a656-412680ba981f
| 2 |
Kinase KIVb phosphorylates numerous nonhistone nuclear proteins as shown by electrophoretic analysis. The addition of kinase KIVb to reaction mixtures containing nonhistone proteins results in the phosphorylation of a spectrum of polypeptides similar to those that are phosphorylated by endogenous nuclear kinases. Nonhistone proteins bound to chromatin appear to be better substrates for KIVb than nonhistones dissociated from chromatin.
|
5e306f9b-61e0-437e-a656-412680ba981f
| 3 |
A comparison of nuclear phosphoproteins phosphorylated either in the intact animal or in vitro (by the addition of kinase KIVb) indicates some differences and some similarities in the patterns of phosphorylation.
|
ad79f7c4-d686-4e64-b10a-016f57e83e94
| 0 |
Purification and some properties of rat liver cysteine oxidase (cysteine dioxygenase). Cysteine oxidase (cysteine dioxygenase, EC 1.13.11.20) was purified approximately 1000-fold from rat liver. The purified enzyme (protein-B) was obtained as an inactive form, which was activated by anaerobic preincubation with L-cysteine.
|
ad79f7c4-d686-4e64-b10a-016f57e83e94
| 1 |
The active form of protein-B was inactivated during aerobic incubation to produce cysteine sulfinate. This inactivation of protein-B was protected by a distinct protein in rat liver cytoplasm, namely stabilizing protein (protein-A). The Ka and Km values for L-cysteine were 0.8-10(-3) M and 1.3
|
ad79f7c4-d686-4e64-b10a-016f57e83e94
| 2 |
-10(-3) M respectively. The enzyme was strongly inhibited by Cu+ and/or Fe2+ chelating agents but not by Cu2+ chelating agent. The optimum pH of enzyme reaction was 8.5-9.5 while that of enzyme activation was 6.8-9.5, with a broad peak.
|
032ead71-c536-47be-81b2-849571c252bd
| 0 |
Conformations of lysine-sensitive aspartokinase. 1. The technique of differential thermal and proteolytic inactivation has been employed as a conformational probe for the lysine-sensitive aspartokinase (EC 2.7.2.4) of Escherichia coli B. 2. L-Amino acid inhibitors of this enzyme each induce a characteristic enzyme conformation. This is evidenced by rates of thermal and proteolytic inactivation and Arrhenius activation energies for thermal inactivation which are characteristic of the amino acid present.
|
032ead71-c536-47be-81b2-849571c252bd
| 1 |
3. Phenylalanine and leucine binding are mutually exclusive as evidenced by competitive behavior in thermal inactivation experiments, suggesting a hydrophobic amino acid binding site with broad specificity. 4. The phenylalanine-dependent conformation and the leucine-dependent conformation differ considerably. In comparison with the native enzyme, the former is more labile to proteolysis by trypsin whereas the latter is more stable.
|
032ead71-c536-47be-81b2-849571c252bd
| 2 |
First-order rate constants for thermal inactivation of the phenylalanine- and leucine-dependent conformations are, respectively, about one-half and one-tenth that of the native enzyme. 5. Items 3 and 4 taken together suggest that the conformations are ligand induced and do not arise via ligand stabilization of spontaneously arising conformers.
|
64f9085a-9cb3-471a-89a8-b99be2998f3b
| 0 |
The sulphatase of ox liver. XIX. On the nature of the polymeric forms of sulphatase A present in dilute solutions. Weight-average elution volumes of sulphatase A (an arylsulphate sulphohydrolase, EC 3.1.6.1) from Sephadex G-200 have been determined as functions of protein concentration, pH, ionic strength and temperature.
|
64f9085a-9cb3-471a-89a8-b99be2998f3b
| 1 |
The results are used to calculate the apparent association equilibrium constants for tetramer formation and the associated standard-state thermodynamic parameters. While the apparent association constant decreased from 10(28) to 10(21) M-3 on increasing the pH from 4.5 to 5.6 at ionic strength 0.1, at any particular pH value studied it was relatively insensitive to temperature variation so that deltaH is close to zero and tetramer formation in solution is associated with a positive entropy change.
|
64f9085a-9cb3-471a-89a8-b99be2998f3b
| 2 |
At pH 5.0, increasing the ionic strength from 0.1 to 2 decreased the association constant by a factor of 100. Methylumbelliferone sulphate has no effect on the association of sulphatase A. The equilibrium results are used to define the degree of association of sulphatase A likely to encountered in experiments designed to elucidate its kinetic properties.
|
64f9085a-9cb3-471a-89a8-b99be2998f3b
| 3 |
In the liver lysosome, the tetramer is probably the dominant species. The monomer and tetramer of sulphatase A have similar, or identical, specific activities with nitrocatechol sulphate and 4-methylumbelliferone sulphate as substrates. With nitrocatechol sulphate, sulphatase A shows Michaelis kinetics under conditions where the monomer is the dominant species and non-Michaelis kinetics where the tetramer is dominant.
|
64f9085a-9cb3-471a-89a8-b99be2998f3b
| 4 |
There is apparently a negative cooperativity between the monomer units in the tetramer. In 2 mM sodium taurodeoxycholate and 0.035 M MnCl2, but not in 0.1 M NaCl, the tetramer shows Michaelis kinetics. This is not due to dissociation of the tetramer. The critical micellar concentration of sodium taurodeoxycholate is about 0.8
|
64f9085a-9cb3-471a-89a8-b99be2998f3b
| 5 |
mM in both 0.1 M NaCl and 0.035 M McCl2 but the aggregation number is greater in the latter.
|
d9cae154-5828-48d1-9f41-effd5fefffa0
| 0 |
Purification and characterization of an extracellular exo-D-galacturonanase of Aspergillus niger. A D-galacturonanase (EC 3.2.1.67) catalyzing the degradation of D-galacturonans by terminal action pattern was purified from a culture filtrate of Aspergillus niger by a procedure including the salting-out with ammonium sulfate, precipitation by ethanol, chromatography on DEAE-cellulose, and gel chromatography on Sephadex G-100.
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d9cae154-5828-48d1-9f41-effd5fefffa0
| 1 |
The obtained preparation was slightly contaminated by an enzymically inactive protein fraction. Maximum activity and stability of the enzyme was observed at pH 5.2. The enzyme degrades digalacturonic acid, p-nitrophenyl-alpha-D-galactopyranuronide, as well as oligogalacturonides containing at the nonreducing end 4-deoxy-L-threo-hexa-4-enopyranosyluronate.
|
d9cae154-5828-48d1-9f41-effd5fefffa0
| 2 |
It differs from all A. niger enzymes so far described which degrade D-galaturonans by the terminal action pattern, in not clearly preferring low-molecular substrates. It is therefore classified as an exo-D-galacturonanase.
|
19304f57-7a57-48c0-8eef-d2f0477bad51
| 0 |
The biosynthesis of multi-L-arginyl-poly(L-aspartic acid) in the filamentous cyanobacterium Anabaena cylindrica. The cyanobacteria produce multi-L-arginyl-poly (aspartic acid), a high molecular weight (Mr=25 000-125 000) branched polypeptide consisting of a poly(aspartic acid) core with L-arginyl residues peptide bonded to each free carboxyl group of the poly(aspartic acid).
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19304f57-7a57-48c0-8eef-d2f0477bad51
| 1 |
An enzyme which will elongate Arg-poly(Asp) has been isolated and purified 92-fold from the filamentous cyanobacterium Anabaena cylindrica. The enzyme incorporates arginine and aspartic acid into Arg-poly(Asp) in a reaction which requires ATP, KCl, MgCl2, and a sulfhydryl reagent. The enzymatic incorporation of arginine is dependent upon the presence of L-aspartic acid but not visa versa, a finding which suggests the order of amino acid addition to the branched polypeptide-aspartic acid is added to the core followed by the attachment of an arginine branch.
|
19304f57-7a57-48c0-8eef-d2f0477bad51
| 2 |
The elongation of Arg-poly(Asp) in-vitro is insensitive to the addition of protein synthesis inhibitors and to the addition of nucleases. These findings support the notion previosly suggested from in-vivo studies that Arg-poly(Asp) is synthesized via a non-ribosomal route and also demonstrate that amino-acetylated transfer-RNAs play no part in at least one step of the biosynthetic mechanism.
|
32651fb7-ce61-405e-9e21-9f7fd7e74826
| 0 |
Separation of subchloroplast membrane particles by counter-current distribution. Counter-current distribution in an aqueous Dextran-polyethylene glycol two-phase system has been used to fractionate membrane fragments obtained by press treatment of Class II chloroplasts. By the counter-current distribution technique membrane particles are separated according to their surface properties such as charge and hydrophobicity.
|
32651fb7-ce61-405e-9e21-9f7fd7e74826
| 1 |
The fractions obtained were analysed with respect to photochemical activities, chlorophyll and P-700 contents. The Photosystem II enrichment after counter-current distribution was better than that obtained by differential centrifugation of the disrupted chloroplasts. However, the best separation of Photosystem I and II enriched particles could be achieved if differential centrifugation was combined with the counter-current distribution technique.
|
32651fb7-ce61-405e-9e21-9f7fd7e74826
| 2 |
Each centrifugal fraction could be further separated into Photosystems I and II enriched fractions since the Photosystem II particles preferred the dextran-rich bottom phase while the Photosystem I particles preferred the polyethylene glycol-rich top phase. By this procedure it was possible, without the use of detergents, to obtain vesicles which were more enriched in Photosystem II as compared to intact grana stacks.
|
32651fb7-ce61-405e-9e21-9f7fd7e74826
| 3 |
The partition behaviour of undisrupted Class II chloroplasts and the Photosystem I centrifugal fraction was the same. This similarity indicated that the membrane which is exposed to the surrounding polymers by the Class II chloroplasts is the Photosystem I rich membrane of the stroma lamellae.
|
97b7bfc2-c696-4a84-aef6-68491d79eb86
| 0 |
Stimulation of ATP synthesis in Halobacterium halobium R1 by light-induced or artifically created proton electrochemical potential gradients across the cell membrane. The relationship between proton movement and phosphorylation in Halo-bacterium halobium R1 has been investigated under anaerobic conditions. The light-induced changes in the bacteriorhodopsin are accompanied by proton movements across the membrane which result in pH changes in the suspending medium.
|
97b7bfc2-c696-4a84-aef6-68491d79eb86
| 1 |
The initial alkaline shift is shown to be closely paralleled by (and hence correlated with) ATP synthesis. Acidification of the medium in the presence of valinomycin, under conditions of low external potassium, brings about ATP synthesis in the dark.
|
84af6541-c4cd-479b-b4bd-27daecfa1630
| 0 |
Mechanism of active shrinkage in mitochondria. II. Coupling between strong electrolyte fluxes. 1. Addition of succinate to valinomycin-treated mitochondria incubated in KCL causes a large electrolyte penetration. The process depends on a steady supply of energy and involves a continuous net extrusion of protons. Rates of respiration and of electrolyte penetration proceed in a parallel manner.
|
84af6541-c4cd-479b-b4bd-27daecfa1630
| 1 |
2. A passive penetration of K+ salt of permeant anions occurs in respiratory-inhibited mitochondria after addition of valinomycin. Addition of succinate at the end of the passive swelling starts an active extrusion of anions and cations with restoration of the initial volume. The shrinkage is accompanied by a slow reuptake of protons.
|
84af6541-c4cd-479b-b4bd-27daecfa1630
| 2 |
The initiation of the active shrinkage correlates with the degree of stretching of the inner membrane. The extrusion of electrolytes is inhibited by nigericin, while it is only slightly sensitive to variations of the valinomycin concentration larger than two orders of magnitude. 3. Passive swelling and active shrinkage occurs also when K+ is replaced by a large variety of organic cations.
|
84af6541-c4cd-479b-b4bd-27daecfa1630
| 3 |
The rate of organic cation penetration is enhanced by tetraphenylboron, while the rate of electrolyte extrusion is insensitive to variation of the tetraphenylboron concentration. 4. Active shrinkage, either with K+ or organic cation salts, is inhibited by weak acids. The phosphate inhibition is removed by SH inhibitors.
|
84af6541-c4cd-479b-b4bd-27daecfa1630
| 4 |
The active shrinkage is also inhibited by mersalyl to an extent of about 60%. 5. Three models of active shrinkage are discussed: (a) mechanoprotein, (b) electrogenic proton pump, and (c) proton-driven cation anion pump.
|
35b7dc55-8250-4311-9107-6106f8e0e925
| 0 |
Inhibitory effect of p-nitrothiophenol in the light on the photosystem II activity of spinach chloroplasts. The treatment of spinach chloroplasts with p-nitrothiophenol in the light at acidic and neutral pH'S caused specific inhibition of the Photosystem II activity, whereas the same treatment in the dark did not affect the activity at all.
|
35b7dc55-8250-4311-9107-6106f8e0e925
| 1 |
The photosystem I activity was not inhibited by p-nitrothiophenol both in the light and in the dark. The inhibition was accompanied by changes of fluorescence from chloroplasts. As observed at room temperature, the 685-nm band was lowered by the p-nitrothiophenol treatment in the light and, at liquid nitrogen temperature, the relative height of the 695-nm band to the 685-nm band increased and the 695-nm band shifted to longer wavelengths.
|
35b7dc55-8250-4311-9107-6106f8e0e925
| 2 |
The action spectra for these effects of p-nitrothiophenol on the activity and fluorescence showed a peak at 670 nm with a red drop at longer wavelengths. It was concluded that the light absorbed by Photosystem II is responsible for the chemical modification of chloroplasts with p-nitrothiopehnol to causing the specific inhibition of Photosystem II.
|
d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 0 |
Relations between the electrical potential, pH gradient, proton flux and phosphorylation in the photosynthetic membrane. The transmembrane electrical potential (deltaphi), the proton flux (H+), the rate of electron transport (e), the pH gradient (deltapH) and the rate of phosphorylation (ATP) were measured in chloroplasts of spinach.
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d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 1 |
Photosynthesis was excited periodically with flashes of variable frequencies and intensities. A new method is described for determining the rate of electron transport and proton flux. Under conditions where the rate of electron transport and proton flux are not pH controlled the following correlations were found in the range 50 mV less than or equal to deltaphi less than or equal to 125 mV and 1.8
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d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 2 |
less than or equal to deltapH less than or equal to 2.7: (1) The pH gradient, deltapH, increases with H+ independently of Phout between 7-9. (2) The rate of phosphorylation, ATP, depends exponentially on deltapH (at constant deltaphi) and is independent of pHout between 7-9.
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d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 3 |
(3) The rate of phosphorylation, ATP, depends also on deltaphi (at constant deltapH and at constant proton flux H+). (4) The proton flux via the ATPase pathway, Hp+, depends non-linearly on the ratio of the proton concentrations: Hp+ approximately (Hin+/Hout+)b, (b=2.3
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d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 4 |
--2.6). The proton flux via the basal pathway, Hb+, depends linearly on the ratio of the proton concentrations: Hb+ approximately (Hin/Hout). (5) The ratio deltaH+/ATP (e/ATP, i.e. the ratio of the total proton flux, Hp+ + Hb+, and the rate of ATP formation, ATP, depends strongly on deltaphi and on deltapH.
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d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 5 |
The ratio is deltaH+/ATP approximately 3 (e/ATP approximately 1.5) at deltapH 2.7 and deltaphi = 125 mV. (6) It is supposed that the reason for the dependence of deltaH+/ATP on deltaphi anddeltapH is the different functional dependence of the basal proton flux Hb+ and the phosphorylating proton flux Hp+ on deltapH and deltaphi.
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d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 6 |
The calculation of deltaH+/ATP on the basis of this assumption is in fair agreement with the experimental values. Also the "threshold" effects can be explained in this way. (7) The ratio of deltaHp+/ATP, i.e. the ratio of the phosphorylating proton flux Hp+ and ATP, is deltaHp+/ATP APPROXIMATELY 2.4
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d0dbd025-f0fa-4fda-8d5a-eecdba2669ff
| 7 |
.
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6c9757cf-e414-4820-b288-2035531d243f
| 0 |
A possible mechanism of the generation of singlet molecular oxygen in nadph-dependent microsomal lipid peroxidation. A simplified system, consisting of NADPH, Fe3+-ADP, EDTA, liposomes, NADPH-cytochrome c reductase and Tris - HCl buffer (pH 6.8), has been employed in studies of the generation of singlet oxygen in NADPH-dependent microsomal lipid peroxidation.
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6c9757cf-e414-4820-b288-2035531d243f
| 1 |
The light emitted by the system involves 1deltag type molecular oxygen identifiable by its characteristic emission spectrum and its behavior with beta-carotene. The generation of another excited species (a compound in the triplet state) could be demonstrated in this system by changes of light intensity and emission spectra which arise from photosensitizer (9,10-dibromoanthracene sulfonate, eosin, Rose-Bengal)-mediated energy transfers.
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6c9757cf-e414-4820-b288-2035531d243f
| 2 |
Chemiluminescence in the visible region was markedly quenched by various radical trappers and by an inhibitor of NADPH-cytochrome c reductase, but not by superoxide dismutase. During the early stage of lipid peroxidation, the intensity of chemiluminescence was proportional to the square of the concentration of lipid peroxide.
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6c9757cf-e414-4820-b288-2035531d243f
| 3 |
These characteristics suggest that singlet oxygen and a compound in the triplet state (probably a carbonyl compound) are generated by a self-reaction of lipid peroxy radicals.
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f5fd8656-0ce8-4b5a-9b57-ac84311a859a
| 0 |
Primary reactions of photosystem II at low pH. I. Prompt and delayed fluorescence. Prompt and delayed chlorophyll fluorescence have been studied in broken spinach chloroplasts at pH values down to 2.6. No direct effect of low pH on the primary charge separation in Photosystem II was observed. The irreversible inactivation of a secondary electron donor in a narrow pH range around pH 4.5
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f5fd8656-0ce8-4b5a-9b57-ac84311a859a
| 1 |
was demonstrated. At lower pH values the photooxidized form of a more primary electron donor, revealed by its efficient fluorescence quenching, was reduced with a half time of about 200 mus, 25% by another electron donor and 75% by back reaction with the reduced acceptor. The electron donation had a half time of 800 mus and was practically irreversible.
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f5fd8656-0ce8-4b5a-9b57-ac84311a859a
| 2 |
The back reaction had a pH dependent half time: about 270 mus at pH 4 and increasing towards lower pH. The competition of both reactions resulted in a net efficiency of the charge separation at pH 4 of 25%, increasing towards lower pH.
|
feaea3a1-0727-473e-aa21-b45f9c499359
| 0 |
Immunological similarity between NADH-cytochrome b5 reductase of erythrocytes and liver microsomes. In a number of animal species soluble NADH-cytochrome b5 reductase of erythrocytes was compared with membrane-bound NADH-cytochrome b5 reductase of liver microsomes by using an antibody to purified NADH-cytochrome b5 reductase from rat liver microsomes. The results obtained indicated clearly that they are immunologically very similar to each other. The data with erythrocyte ghosts suggested that cytochrome b5 and NADH-cytochrome b5 reductase are also present in the ghost.
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e6764b5c-acab-4c95-87dc-e58ca0e50841
| 0 |
Effect of NADP on light-induced cytochrome changes in membrane fragments from a blue-green alga. The effect of NADP+ on light-induced steady-state redox changes of membrane-bound cytochromes was investigated in membrane fragements prepared from the blue-green algae Nostoc muscorum (Strain 7119) that had high rates of electron transport from water to NADP+ and from an artificial electron donor, reduced dichlorophenolindophenol (DCIPH2) to NDAP+.
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e6764b5c-acab-4c95-87dc-e58ca0e50841
| 1 |
The membrane fragments contained very little phycocyanin and had excellent optical properties for spectrophotometric assays. With DCIPH2 as the electron donor, NADP+ had no effect on the light-induced redox changes of cytochromes: with or without NADP+, 715- or 664-nm illumination resulted mainly in the oxidation of cytochrome f and of other component(s) which may include a c-type cytochrome with an alpha peak at 549nm.
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e6764b5c-acab-4c95-87dc-e58ca0e50841
| 2 |
With 664 nm illumination and water as the electron donor, NADP+ had a pronounced effect on the redox state of cytochromes, causing a shift toward oxidation of a component with a peak at 549 nm (possibly a c-type cytochrome), cytochrome f, and particularly cytochrome b559.
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e6764b5c-acab-4c95-87dc-e58ca0e50841
| 3 |
Cytochrome b559 appeared to be a component of the main noncyclic electron transport chain and was photooxidized at physiological temperatures by Photosystem II. This photooxidation was apparent only in the presence of a terminal acceptor (NADP+) for the electron flow from water.
|
68017c70-becf-42d2-8d01-e959e35d1d38
| 0 |
Kinetic studies on cytochrome c oxidase by combined epr and reflectance spectroscopy after rapid freezing. 1. Techniques and experiments are described concerned with the millisecond kinetics of EPT-detectable changes brought about in cytochrome c oxidase by reduced cytochrome c and, after reduction with various agents, by reoxidation with O2 or ferricyanide.
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68017c70-becf-42d2-8d01-e959e35d1d38
| 1 |
Some experiments in the presence of ligands are also reported. Light absorption was monitored by low-temperature reflectance spectroscopy. 2. In the rapid phase of reduction of cytochrome c oxidase by cytochrome c (less than 50 ms) approx. 0.5 electron equivalent per heme a is transferred mainly to the low-spin heme component of cytochrome c oxidase and partly to the EPR-detectable copper.
|
68017c70-becf-42d2-8d01-e959e35d1d38
| 2 |
In a slow phase (less than 1 s) the copper is reoxidized and high-spin ferric heme signals appear with a predominant rhombic component. Simultaneously the absorption band at 655 nm decreases and the Soret band at 444 nm appears between the split Soret band (442 and 447 nm) of reduced cytochrome a.
|
68017c70-becf-42d2-8d01-e959e35d1d38
| 3 |
3. On reoxidation of reduced enzyme by oxygen all EPR and optical features are restored within 6 ms. On reoxidation by O2 in the presence of an excess of reduced cytochrome c, states can be observed where the low-spin heme and copper signals are largely absent but the absorption at 655 nm is maximal, indicating that the low-spin heme and copper components are at the substrate side and the component(s) represented in the 655 nm absorption at the O2 side of the system.
|
68017c70-becf-42d2-8d01-e959e35d1d38
| 4 |
On reoxidation with ferricyanide the 655 nm absorption is not readily restored but a ferric high-spin heme, represented by a strong rhombic signal, accumulates. 4. On reoxidation of partly reduced enzyme by oxygen, the rhombic high-spin signals disappear within 6 ms., whereas the axial signals disappear more slowly, indicating that these species are not in rapid equilibrium.
|
68017c70-becf-42d2-8d01-e959e35d1d38
| 5 |
Similar observations are made when partly reduced enzyme is mixed with CO. 5. The results of this and the accompanying paper are discussed and on this basis an assignment of the major EPR signals and of the 655 nm absorption is proposed, which in essence is that published previously (Hartzell, C.
|
68017c70-becf-42d2-8d01-e959e35d1d38
| 6 |
R., Hansen, R.E. and Beinert, H. (1973) Proc. Natl. Acad. Sci. U.S. 70, 2477-2481). Both the low-spin (g=o; 2.2; 1.5) and slowly appearing high-spin (g=6; 2) signals are attributed to ferric cytochrome a, whereas the 655 nm absorption is thought to arise from ferric cytochrome a3, when it is present in a state of interaction with EPR-undectectable copper.
|
68017c70-becf-42d2-8d01-e959e35d1d38
| 7 |
Alternative possibilities and possible inconsistencies with this proposal are discussed.
|
67f6487a-c5f8-4670-8d61-66fccc7271a0
| 0 |
Some thermodynamic and kinetic properties of the primary photochemical reactants in a complex from a green photosynthetic bacterium. We have examined the bacteriochlorophyll reaction-center complex of Chlorobium limicola f. thiosulfatophilum, strain Tassajara. Our results indicate that the midpoint potential of the primary electron donor bacteriochlorophyll of the reaction center is +250 mV at pH 6.8
|
67f6487a-c5f8-4670-8d61-66fccc7271a0
| 1 |
, while that of cytochrome c-553 is +165 mV. There are two cytochrome c-553 hemes per reaction center, and the light-induced oxidation of each is biphasic (t1/2 of less than 5 mus and approximately 50 mus). We belive that this indicates a two state equilibrium with each cytochrome heme being either close to, or a little removed from, the reaction-center bacteriochlorophyll.
|
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