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PubMed1989

Relative contributions of protein sulfhydryl loss and lipid peroxidation to allyl alcohol-induced cytotoxicity in isolated rat hepatocytes.

The time course of allyl alcohol-induced toxicity was studied in freshly isolated rat hepatocytes. The sequence of events was as follows: an initial, rapid depletion of glutathione (GSH), a subsequent increase in malondialdehyde (MDA) and decrease in protein sulfhydryl groups (PSH), and the eventual loss of membrane integrity. The sulfhydryl compounds, N-acetylcysteine and dithiothreitol, markedly delayed the depletion of GSH, prevented significant loss of PSH, and protected the cells against viability loss. In contrast, the antioxidants, butylated hydroxytoluene and Trolox C, and the iron-chelating agent, deferoxamine, suppressed allyl alcohol-induced MDA production without affecting the depletion of cellular thiols or the loss of viability. The results suggest that the inactivation of protein thiol groups is critical for allyl alcohol toxicity whereas lipid peroxidation is not essential to the toxic process.

1-Propanol↗
PubMed1999

Alteration of free radical metabolism in the brain of mice infected with scrapie agent.

Alteration of free radical metabolism in the mouse brain by scrapie infection was evaluated. The infection of mice with scrapie agent, 87V strain, slightly increased the activities of catalase and glutathione-S-transferase, while it had no effect on glutathione peroxidase, glutathione reductase, and Cu, Zn-superoxide dismutase. Results show that the scrapie infection decreased the activity of mitochondrial Mn-superoxide dismutase by 50% but increased that of monoamine oxidase (p < 0.05). Scrapie infection also increased the rate of mitochondrial superoxide generation (p < 0.05). Following scrapie infection, the level of free-sulfhydryl compounds in brain homogenates slightly decreased, but the content of thiobarbituric-acid-reactive substances and malondialdehyde increased significantly. Electron microscopy indicated that the ultrastructure of mitochondria was destroyed in the brain of scrapie-infected mice. These results suggest that elevated oxygen free radical generation and lowered scavenging activity in mitochondria might cause the free radical damage to the brain. Such deleterious changes in mitochondria may contribute to the development of prion disease.

Animals↗
PubMed1989

Microsomal glutathione-dependent protection against lipid peroxidation acts through a factor other than glutathione peroxidase and glutathione S-transferase in rat liver.

Ascorbate-Fe3+-induced and NADPH-induced lipid peroxidation of rat liver microsomes were inhibited by glutathione (GSH). This inhibition was due to microsomal GSH-dependent factor. This factor was heat labile, and storage of microsomes at 4 degrees C for 1 week diminished the activity. GSH could not be substituted by other sulfhydryl compounds tested. Deoxycholate (1 mM) and bromosulfophthalein (0.1 mM) inhibited GSH-dependent protection but did not inhibit microsomal GSH peroxidase activity. Iodoacetate (10 mM) inhibited GSH-dependent protection but did not inhibit microsomal GSH S-transferase. N-Ethylmaleimide (0.1 mM) and oxidized glutathione (10 mM) inhibited GSH-dependent protection but activated microsomal GSH S-transferase activity. These results indicate the existence of a heat-labile, microsomal GSH-dependent protective factor against lipid peroxidation that acts through a factor other than GSH-peroxidase and GSH S-transferase.

Animals↗
PubMed1975

Beneficial effects of penicillamine treatment on hereditary avian muscular dystrophy.

Hereditary muscular dystrophy in chickens of the New Hampshire strain was treated with penicillamine from the 9th day after hatching to the 425th day. The adult maintenance dose for males was 50 mg/kg per day and for females, 13-65 mg/kg per day. In avian dystrophy, deterioration of the muscle fibers is evidenced in the 2nd mo by an inability of the birds to rise after falling on their backs and by a progressive rigidity of the wings. The drug delayed the onset of symptoms and partially alleviated the debilitating aspects of the disease. Penicillamine produced three major improvements: (a) better righting ability when birds were placed on their backs; (b) greater wing flexibility; (c) and suppression of plasma creatine phosphokinase activity. The results are statistically analyzed and discussed in relationship to Duchenne dystrophy. Normal birds were not affected by penicillamine as judged by these parameters. The rationale for using penicillamine, a sulfhydryl compound with reducing properties, was (a) to attempt to protect essential thiol enzymes in the anabolic and glycolytic pathways against inactivation and (b) to prevent collagen cross-linking and deposition in muscle. Although the precise mechanism of drug action has not been determined. the possible role of penicillamine in mitigating the symptoms of genetic dystrophy in man is under consideration. Further, penicillamine may have a more generalized application i the prevention of contractures in a variety of neuromuscular disorders.

Animals↗
PubMed1991

Pharmacological evidence that captopril possesses an endothelium-mediated component of vasodilation: effect of sulfhydryl groups on endothelium-derived relaxing factor.

Captopril, an angiotensin-converting enzyme inhibitor, reportedly can scavenge superoxide anion (O2-), a property attributed to its sulfhydryl group. The present investigation, using rabbit aortic rings precontracted with either norepinephrine or clonidine, was designed to determine whether captopril possesses an endothelium-dependent component of vasodilation related to its ability to protect endothelium-derived relaxing factor (EDRF) from superoxide-mediated destruction. Also studied were enalaprilat, a nonsulfhydryl angiotensin-converting enzyme-inhibitor, superoxide dismutase, and the sulfhydryl compounds glutathione (GSH), N-2-mercaptopropionylglycine (MPG) and N-acetylcysteine (NAC). Captopril, but not enalaprilat, caused dose-dependent relaxations in preconstricted aortic rings containing an intact endothelium. Rings denuded of endothelium were unresponsive to any dose of captopril. Captopril's vasodilation was not related to prostaglandin influence but was associated with an increase in cyclic GMP. Superoxide dismutase, GSH, MPG and NAC also produced endothelium-dependent relaxations similar to captopril. It was also demonstrated that endothelium-dependent relaxations to acetylcholine were enhanced by captopril, GSH, MPG and NAC but not by enalaprilat. In another set of experiments, the ability of captopril to inhibit superoxide-mediated inactivation of EDRF was examined. Pyrogallol, a potent generator of O2-, and superoxide dismutase, a scavenger of O2-, were used as a basis for comparing a possible scavenging effect of captopril. In preconstricted rings, pyrogallol elicited endothelium-dependent contractions that were attenuated by both captopril and superoxide dismutase. Similar effects were found with GSH, MPG and NAC but not with enalaprilat. These results suggest that captopril's endothelium-dependent vasodilation is due to its sulfhydryl group and the ability of the latter to scavenge O2-, thereby protecting EDRF.

Acetylcholine↗
PubMed1986

Purification and properties of synephrinase from Arthrobacter synephrinum.

Synephrinase, an enzyme catalyzing the conversion of (-)-synephrine into p-hydroxyphenylacetaldehyde and methylamine, was purified to apparent homogeneity from the cell-free extracts of Arthrobacter synephrinum grown on (+/-)-synephrine as the sole source of carbon and nitrogen. A 40-fold purification was sufficient to produce synephrinase that is apparently homogeneous as judged by native polyacrylamide gel electrophoresis and has a specific activity of 1.8 mumol product formed/min/mg protein. Thus, the enzyme is a relatively abundant enzyme, perhaps comprising as much as 2.5% of the total protein. The enzyme essentially required a sulfhydryl compound for its activity. Metal ions like Mg2+, Ca2+, and Mn2+ stimulated the enzyme activity. Metal chelating agents, thiol reagents, denaturing agents, and metal ions like Zn2+, Hg2+, Ag1+, and Cu2+ inhibited synephrinase activity. Apart from (-)-synephrine, the enzyme acted upon (+/-)-octopamine and beta-methoxysynephrine. Molecular oxygen was not utilized during the course of the reaction. The molecular mass of the enzyme as determined by Sephadex G-200 chromatography, was around 156,000. The enzyme was made up of four identical subunits with a molecular mass of 42,000.

Arthrobacter↗
PubMed1997

Glutathione alters the mode of calcium-mediated regulation of adenylyl cyclase in membranes from mouse brain.

We examined the effect of sulfhydryl compounds on the regulation of adenylyl cyclase by calcium in mouse cerebrum membranes. Isoproterenol-stimulated adenylyl cyclase (IP-AC) activity in the membranes was increased by addition of the optimum concentrations of calcium/calmodulin. However, in the presence of 0.01-0.07 mM glutathione (GSH), calcium/calmodulin inhibited the activity. At high concentrations of GSH (1-10 mM), the IP-AC activity was stimulated by calcium/calmodulin to a greater extent than that in the control (no GSH). Cysteine at less than 1.7 mM induced a similar inhibition of the IP-AC activity, but dithiothreitol did not. The activity of IP-AC measured in the absence of calmodulin decreased when calcium levels were greater than 300 microM. GSH at 0.05 mM enhanced the calcium-dependent inhibition (22% inhibition by 200 microM calcium), while 10 mM GSH lowered it. Calmodulin itself had no significant effect on the IP-AC activity, irrespective of the concentrations of GSH involved. It caused a small increase in the IP-AC activity that had been reduced by the presence of calcium and GSH. These results indicate that the redox status of sulfhydryls in adenylyl cyclase plays an important role in the calcium-mediated regulation of the enzyme. The enzyme becomes much more sensitive to the calcium-dependent inhibition after partial reduction of the sulfhydryls via the particular mode of reactin.

Adenylyl Cyclase Inhibitors↗
PubMed1977

Hydrogenase activity and the H2-fumarate electron transport system in Bacteroides fragilis.

Hydrogenase activity and the H(2)-fumarate electron transport system in a carbohydrate-fermenting obligate anaerobe, Bacteroides fragilis, were investigated. In both whole cells and cell extracts, hydrogenase activity was demonstrated with methylene blue, benzyl viologen, flavin mononucleotide, or flavin adenine dinucleotide as the electron acceptor. A catalytic quantity of benzyl viologen or ferredoxin from Clostridium pasteurianum was required to reduce nicotinamide adenine dinucleotide or nicotinamide adenine dinucleotide phosphate with H(2). Much of the hydrogenase activity appeared to be associated with the soluble fraction of the cell. Fumarate reduction to succinate by H(2) was demonstrable in cell extracts only in the presence of a catalytic quantity of benzyl viologen, flavin mononucleotide, flavin adenine dinucleotide, or ferredoxin from C. pasteurianum. Sulfhydryl compounds were not required for fumarate reduction by H(2), but mercaptoethanol and dithiothreitol appeared to stimulate this activity by 59 and 61%, respectively. Inhibition of fumarate reduction by acriflavin, rotenone, 2-heptyl-4-hydroxyquinoline-N-oxide, and antimycin A suggest the involvement of a flavoprotein, a quinone, and cytochrome b in the reduction of fumarate to succinate. The involvement of a quinone in fumarate reduction is also apparent from the inhibition of fumarate reduction by H(2) when cell extracts were irradiated with ultraviolet light. Based on the evidence obtained, a possible scheme for the flow of electrons from H(2) to fumarate in B. fragilis is proposed.

Acriflavine↗
PubMed1983

Inactivation of the beta-adrenergic receptor in skeletal muscle by dithiols.

The effect of sulfhydryl compounds on binding of the beta-adrenergic antagonist (-)-[3H]dihydroalprenolol [(-)-[3H]DHA] to a microsomal fraction from rabbit skeletal muscle was examined. Inhibition of binding by a variety of adrenergic agonists and antagonists and the effects of these agents on adenylate cyclase were consistent with the beta-adrenergic receptor in this tissue being of the beta 2-subtype. Binding of (-)-[3H]DHA was reduced by incubating the membranes with dithiols such as dithiothreitol (DTT), 1,3-dimercapto-2-propanol and 1,4-dimercaptobutane; monothiols were much less potent. DTT-induced decline in (-)-[3H]DHA binding resulted primarily from a decrease in receptor number. Inactivation was partially reversed by the oxidant H2O2. Binding sites could be locked in the inactivated state by incubating DTT-treated membranes with the alkylating agent iodoacetamide. Both beta-adrenergic agonists and antagonists protected against inactivation. Adenylate cyclase activity in the membranes was increased by DTT. The enzyme was rapidly inactivated by H2O2, and this could be partially reversed by DTT. It is concluded that the beta-adrenergic receptor of skeletal muscle contains an essential disulfide moiety which can be inactivated by reducing dithiols. Adenylate cyclase, on the other hand, contains at least one essential sulfhydryl which is preserved by dithiols.

Adenylyl Cyclases↗
PubMed1986

Characterization of carbon disulfide binding in blood and to other biological substances.

Free and bound forms of CS2 are present in subjects exposed to CS2. In rats exposed to 2 mg/liter (approximately 640 ppm) of CS2 for 4 hr, concentrations of acid-labile CS2 (AL CS2, a form of bound CS2 that can be recovered from biological samples by acid treatment) in plasma and red blood cells (RBCs) increased linearly with exposure time. The majority (90%) of the blood AL CS2 was present in the RBCs. About 95% of the AL CS2 in plasma and in RBCs of exposed rats was found in the precipitates after treatment with ammonium sulfate. Incubation of fractionated human RBC lysates with CS2 showed that CS2 binding in these fractions was proportional to the hemoglobin concentration. These observations show that in blood, CS2 binds (in the form of AL CS2) mainly to hemoglobin and to a small extent to other blood proteins. Binding of CS2 to small molecules, including amino acids, accounted for only a small fraction of blood AL CS2. In in vitro studies, CS2 also bound to human albumin, gamma-globulin, and horse heart myoglobin. It was also found that CS2 binds to amino and sulfhydryl compounds at physiological pH. Plasma incubated with CS2 was found to chelate copper. Chelation of copper-containing enzyme by the reaction products of CS2 and biological amines has been observed and has been proposed as one of the mechanisms by which CS2 induces neurotoxicity (M.J. McKenna and V. DiStefano, 1977b, J. Pharmacol. Exp. Ther. 202, 253-266). Radioisotope studies showed that a substantial portion of the radioactivity could not be released from 14CS2-treated plasma and serum upon acid treatment at elevated temperature. These studies suggest the existence of non-acid-labile bound CS2, besides AL CS2, in plasma and serum treated with CS2.

Amino Acids↗
PubMed1982

Factors which affect DNA strand breakage in human leukocytes exposed to a tumor promoter, phorbol myristate acetate.

We have recently reported that phorbol myristate acetate (PMA) induces extensive DNA strand break damage in human peripheral blood leukocytes. The mechanism of action involves superoxide anion and hydrogen peroxide which are generated by phagocytes during the "respiratory burst." In this report, we describe the effect of various inhibitors and scavengers on PMA-induced DNA damage. Azide and cyanide greatly increased the level of damage; sulfhydryl compounds (glutathione, cysteine, and cysteamine) and ascorbate markedly decreased the level of damage. Hydroxyl radical scavengers such as dimethyl sulfoxide (DMSO) and glycerol also decreased the level of damage but apparently did so by inhibiting the respiratory burst. Diethyldithiocarbamate (DDC) increased the level of DNA damage at low concentrations (less than 1 mM), but decreased DNA damage at greater than or equal to 1 mM. The results are consistent with a mechanism involving superoxide anion and hydrogen peroxide, but the precise reaction (free radical or enzymatic) responsible for DNA strand breakage has not been determined. The PMA-stimulated phagocyte is an interesting model system for looking at "active oxygen" mediated DNA damage and factors which influence it.

Ascorbic Acid↗
PubMed2001

Effects of 2(RS)-n-propylthiazolidine-4(R)-carboxylic acid on extrahepatic sulfhydryl levels in mice treated with acetaminophen.

The cysteine (Cys) precursor 2(RS)-n-propylthiazolidine-4(R)-carboxylic acid (PTCA) has been shown to protect against acetaminophen (APAP)-induced hepatic GSH, GSSG, and Cys depletion and hepatic necrosis. The aim of this study was to determine the effects of PTCA on the concentrations of sulfhydryl compounds in extrahepatic tissues, including renal cortex, whole blood, and brain, in C57BL/6 mice treated with hepatotoxic doses of APAP. PTCA (1-5 mmol/kg, i.p.) was administered 30 min after the administration of APAP at a dose (800 mg/kg; 5.29 mmol/kg, i.p.) that depleted hepatic GSH and Cys at 4 hr by 95 and 86%, respectively. Tissue concentrations of GSH and Cys were determined by HPLC. At 4 hr following APAP administration, renal cortical GSH and Cys concentrations were decreased to 64 and 39%, respectively, of vehicle-treated control values, and blood concentrations were decreased to 87 and 30%, respectively, of vehicle controls. Brain GSH and Cys were not depleted by APAP. PTCA at 5 mmol/kg (i) attenuated the APAP-induced depletion of GSH and Cys at 4 hr in renal cortex (78 and 65%, respectively, of vehicle controls), (ii) prevented APAP-induced Cys depletion in blood (670% of vehicle controls) with no effect on GSH concentration (94% of vehicle controls), and (iii) increased GSH and Cys concentrations in brain (119 and 411%, respectively, of vehicle controls). The results demonstrate a high degree of tissue selectivity in the APAP-induced depletion of GSH and Cys, and in the effectiveness of PTCA in maintaining and even elevating sulfhydryl levels in extrahepatic tissues of APAP-treated mice.

Acetaminophen↗
PubMed2002

Changes in antioxidant status of heart muscle tissue in experimental diabetes in rabbits.

The present study was designed to evaluate the oxidative stress-related parameters in alloxan-induced diabetes in rabbits. After 3, 6, 12 and 24 weeks of hyperglycaemia the enzymatic and non-enzymatic factors were measured in heart tissue of diabetic and control groups. Superoxide dismutase and glutathione peroxidase activities and the contents of total sulfhydryl compounds significantly increased at all time intervals. Catalase activity increased initially (after 3 and 6 weeks), decreased after 12 weeks and increased again at the 24th week of the experiment. Glutathione reductase activity increased initially (at 3rd week), decreased below control level after 6 and 12 weeks, then increased again. Ascorbic acid concentration decreased after 3 and 6 weeks, and increased at the 12th and 24th weeks. The level of lipid peroxidation products was reduced after 3, 6 and 12 weeks of the experiment. After 24 weeks it was significantly elevated. These data suggest that hyperglycaemia induces oxidative stress in the heart but the defense mechanisms in the heart tissue are fairly efficacious against oxidative injury.

Alloxan↗
PubMed1992

Hemorrhagic, fibrinogenolytic and edema-forming activities of the venom of the colubrid snake Philodryas olfersii (green snake).

The venom of P. olfersii has high hemorrhagic, edema-inducing and fibrin(ogen)olytic activities. It is devoid of thrombin-like, procoagulant, phospholipase A2 and platelet aggregating enzymes. The main activities are metalloproteinases inhibited by metal chelators (EDTA and 1,10-phenanthroline) and sulfhydryl compounds (DTT and cysteine). The hemorrhagic and fibrinogenolytic enzymes were partially purified by gel filtration on HPLC. The hemorrhagic activity of the venom was neutralized by commercial horse antivenoms to Bothrops species, as well as by rabbit antisera specific for hemorrhagic factors isolated from these Bothrops venoms. No immunoprecipitin reactions were obtained, indicating that the few epitopes of the P. olfersii hemorrhagin are involved in these neutralization reactions. The fibrinogenolytic enzyme cleaves A alpha-chain more quickly than the B beta-chain of human fibrinogen. The venom also solubilizes fibrin. This solubilization appears to occur from the hydrolysis of unpolymerized alpha-chain and cross-linked gamma-gamma dimer. The fibrin peptide products are distinct from those produced by plasmin.

Animals↗
PubMed2001

Differences in antioxidant status in skeletal muscle tissue in experimental diabetes.

BACKGROUND: It has been suggested that oxidative stress may play an important role in pathogenesis of diabetic complications. The present study was designed to evaluate the oxidative stress-related parameters in alloxan (A)-induced long-term diabetes in rabbits. METHODS: After 3, 6 and 12 weeks of diabetes, activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutathione reductase (GSSG-R) and concentrations of ascorbic acid (AA) and free sulfhydryl compounds (SH) were measured in skeletal muscle of diabetic rabbits and the normal control subjects. The products of lipid peroxidation (MDA) were also estimated. RESULTS: In our tests, the muscle SOD activity, SH and AA concentrations were significantly reduced. CAT activity increased significantly at all time intervals. GSH-Px activity decreased after 3 weeks and then remained at the control level. GSSG-R activity decreased progressively at 3rd and 6th week and then significantly increased. MDA level increased initially, dropped below baseline after 6 weeks and then remained at the level of the control group. CONCLUSIONS: The changes observed in the present experiment suggest a significant imbalance in antioxidative system in the skeletal muscle of rabbits with alloxan-induced diabetes. Such study may lead to therapeutic approaches for limiting the damage from oxidation reactions and preventing the diabetic complications.

Animals↗
PubMed1978

The reaction of mercaptans with tyrosinases and hemocyanins.

1. Titration of Neurospora tyrosinase with 2-mercaptoethanol shows that the increase of absorbance at 700 nm is directly correlated to the loss of enzymatic activity. Approximately 2 mol of 2-mercaptoethanol per mole of protein are needed for full development of the green, enzymatically inactive complex. The increase of absorbance at 700 nm is also proportional to the intensity of the EPR signal and the amount of non-covalently bound 2-[35S] mercaptoethanol to the enzyme. The maximal EPR intensity reaches 70% of the protein concentration and at most 0.7--0.8 mol of 2-[35S] mercaptoethanol is bound per mol of enzyme. 2. Stopped-flow measurements show that in the reaction between 2-mercaptoethanol and Neurospora tyrosinase a raction intermediate with a strong absorption band at 360 nm is formed in an apparent second-order reaction. This intermediate displays no EPR-detectable signals. The intermediate decays in a similar complex fashion as the absorption band at 700 nm is formed. 3. The reaction of Neurospora tyrosinase with a variety of sulfhydryl compounds was also investigated. In most cases green coloured, enzymatically inactive complexes are formed displaying slightly different EPR signals. However, with cysteine and cysteamine violet coloured, enzymatically inactive complexes are formed which show rather different EPR signals. The integrated EPR intensities amount to 40--70% of the protein concentration. Based on simulations of 9 and 35 GHz spectra all observed EPR spectra can be represented as true S = 1/2 systems. The cysteamine complex can be interpreted as arising from a mixed valence Cu2+ . Cu+ complex. The 2-mercaptoethanol spectra can, however, arise from sulphur radicals. 4. Treatment of Agaricus bispora tyrosinase and Cancer pagures hemocyanin with 2-mercaptoethanol results in green-coloured, EPR detectable complexes similar to the one found with Neurospora tyrosinase. No such complexes are formed when hemocyanins from Helix pomatia and Panulirus interruptus were treated with this reagent.

Basidiomycota↗
PubMed1982

Biochemical studies of tick embryogenesis. V. Purification and partial characterization of isocitrate lyase from eggs of the tick Hyalomma dromedarii.

1. Isocitrate lyase (Ds-isocitrate glyoxylate-lyase, EC 4.1.3.1) was purified approximately 93-fold from developing embryos of the tick Hyalomma dromedarii. 2. The enzyme requires Mg2+ (Km 2.1 mM) and sulfhydryl compounds for maximal activity and has a pH optimum of 7.4 in phosphate buffer. The Km of the enzyme for isocitrate is 2.4 mM. 3. Data obtained from the pH effect on Km implicate the presence of at least three dissociable groups with pK's of 5.8, 6.8 and 7.4 involved in the enzyme catalysis. 4. At the optimal pH the enzyme is competitively inhibited by oxaloacetate (Ki 0.7 mM) and pyruvate (Ki 0.63 M), noncompetitively inhibited by acetyl-CoA (Ki 1.6 mM) and succinate (Ki 1.35). 5. Inhibition by phosphoenolpyruvate in pH-dependent. The enzyme is noncompetitively inhibited by phosphoenolpyruvate at pH 4.4 (Ki 1.33 mM), 5.6 (Ki 1.7 mM) and pH 8 (Ki 1.36 mM), and competitively inhibited at pH 6.5 (Ki 1.58) and 6.8 (Ki 3.0 mM). 6. The results suggest the regulation of H. dromedarii isocitrate lyase activity during embryonic development by variations in the differential rate of enzyme synthesis an in the intracellular levels of certain metabolites.

Animals↗
PubMed1972

A collagenolytic system produced by primary cultures of rheumatoid nodule tissue.

A collagenase and a neutral protease have been insolated and characterized from primary cultures obtained from rheumatoid subcutaneous nodules. Release of both active enzymes was maximal between the 3rd and 7th days of culture and was stimulated by the presence of small amounts of colchicine (0.1 mug/ml) added to the culture medium. Both the protease and the collagenase from nodule tissue were active at physiologic pH and were inhibited by chelating agents, sulfhydryl compounds, and 1:40 dilutions of human serum. Both enzymes appeared to have a molcular size equivalent to similar enzymes found in cultures of rheumatoid synovium. The nodule collagenase was purified by chromatography on molecular sieve columns followed by affinity chromatography. The pure enzyme cleaved collagen in solution at 24 degrees C at the locus common for mammalian collagenases to act: three quarters of the distance from the amino-terminus. Under the same conditions the purified enzyme cleaved gelatin (denatured collagen) at the same locus. It is likely therefore that the collagenase in rheumatoid connective tissues functions to produce the initial cleavage of collagen and that after the initial reaction products are denatured, proteases digest them into smaller polypeptides more rapidly than does the collagenase itself. Since rheumatoid nodules grow centrifugally at the expense of the palisading fibroblast layer it seems possible that the central necrotic areas are caused by release of collagenase and protease from the highly cellular palisading zone resulting in the destruction of the extracellular collagen matrix.

Arthritis, Rheumatoid↗