Showing papers on "Proteolytic enzymes published in 1969"
TL;DR: The hypothesis is formulated that most of the xanthine oxidase of rat liver supernatant is a dehydrogenase (Type D), and may be converted (activated) into an oxid enzyme (Type O).
694 citations
TL;DR: It is established that the microfibrils consist of a different connective tissue protein (or proteins) that is neither collagen nor elastin during embryologic development, and may play a primary role in the morphogenesis of the elastic fiber.
Abstract: The two morphologically different constituents of the mature elastic fiber, the central amorphous and the peripheral microfibrillar components, have been separated and partially characterized. A pure preparation of elastic fibers was obtained from fetal bovine ligamentum nuchae by extraction of the homogenized ligament with 5 M guanidine followed by digestion with collagenase. The resultant preparation consisted of elastic fibers which were morphologically identical with those seen in vivo. The microfibrillar components of these elastic fibers were removed either by proteolytic enzymes or by reduction of disulfide bonds with dithioerythritol in 5 M guanidine. The microfibrils solubilized by both methods were rich in polar, hydroxy, and sulfur-containing amino acids and contained less glycine, valine, and proline than the amorphous component of the elastic fiber. In contrast, the amino acid composition of the amorphous component was identical with that previously described for elastin. This component demonstrated selective susceptibility to elastase digestion, but was relatively resistant to the action of other proteolytic enzymes and to reduction. These observations establish that the microfibrils consist of a different connective tissue protein (or proteins) that is neither collagen nor elastin. During embryologic development the microfibrils form an aggregate structure before the amorphous component is secreted. These microfibrils may therefore play a primary role in the morphogenesis of the elastic fiber.
636 citations
Journal Article•
TL;DR: A tissue antigen reactive with sera from SLE patients has been partially characterized and is an acidic macromolecule which has the electrophoretic mobility of an α 1 globulin and is resistant to most proteolytic enzymes including trypsin, pepsin and chymotrypsin.
Abstract: Summary A tissue antigen reactive with sera from SLE patients has been partially characterized. The antigen is a soluble cytoplasmic component which is present in a variety of human tissues and is distinct from known nuclear antigens. It is an acidic macromolecule which has the electrophoretic mobility of an α 1 globulin and is resistant to most proteolytic enzymes including trypsin, pepsin and chymotrypsin. Antigenicity is destroyed by 0.02 M periodate and by 0.001 M parahydroxy mercuribenzoate. Antibodies to this antigen have been found in 40% of unselected LE sera and were absent from a large number of sera from normal persons and from the sera of patients with other connective tissue disorders.
478 citations
TL;DR: It is suggested that the coat in bloodstream trypanosomes constitutes a replaceable surface which, after being replaced, enables thetrypanosome to escape the effects of host antibodies.
Abstract: Pathogenic trypanosomes in their bloodstream phase have a smooth and compact coat 12-15 nm thick enveloping the entire surface membrane of the body and flagellum. In the sleeping-sickness trypanosome Trypanosoma rhodesiense this coat is absent from the stages of development in the midgut of the tsetse-fly vector and from their counterparts obtained by cultivation of the trypanosome in vitro. In the salivary glands of the vector, however, the coat is reacquired as the trypanosomes transform from epimastigote forms into the metacyclic stage which is infective to the mammalian host. This loss and acquisition of the surface coat can be correlated with the cyclical changes in net surface charge on the trypanosome which have been observed by other workers. The trypanosome populations of successive relapses in the blood are known to differ in their surface antigens (agglutinogens) and the loss of antigenic identity detected when any of these populations are put into culture indicates that these variable antigens are located in the surface coat. It is suggested that the coat in bloodstream trypanosomes constitutes a replaceable surface which, after being replaced, enables the trypanosome to escape the effects of host antibodies. The coat is therefore an adaptation to life in the bloodstream. Reacquisition of the surface coat by the metacyclic trypanosome after development in the vector may reflect reversion to a ‘basic’ antigenic type at this stage, preparatory to invading the blood of the mammalian host. The surface coat may be removed by the wide-spectrum proteolytic enzyme pronase, and this fact together with evidence from pH/mobility relationships and chemical analysis of the variable antigens suggest that the coat is basically proteinaceous. The coat may facilitate pinocytosis by binding proteins at sites within the pocket surrounding the base of the flagellum. In the non-pathogenic trypanosome T. lewisi a more diffuse filamentous coat is present in bloodstream forms and absent from culture forms. This trypanosome is said to carry a negative charge in both bloodstream and culture phases, so it seems likely that the nature of the coat in T. lewisi is different from that found in the pathogenic trypanosomes. In all these trypanosomes the flagellar membrane adheres to the surface membrane of the body throughout the life-cycle. Along the zone of adhesion lies a regular row of junctional complexes of the macula adherens type which, it is argued, serve in attachment. These attachments persist regardless of changes in the intervening cell surfaces.
428 citations
TL;DR: An assay of proteolytic activity is described, with N, N-dimethylcasein or N,N-dim methylhemoglobin as substrate, which is from 10 to several hundred times more sensitive than the standard caseinolytic assay of Kunitz.
275 citations
TL;DR: It is proposed that the control of the diglyceride concentration of locust haemolymph during flight is mediated, at least in part, by a peptide hormone released from the corpora cardiaca.
268 citations
TL;DR: The findings indicate that, in addition to causing acute illness, inhalation of this material may lead to irreversible impairment of lung function, and that insidious lung damage could occur without episodes of overt illness.
247 citations
TL;DR: After extraction of the torpedo electric organ, three main molecular species, A, C, D, carrying the acetylcholinesterase activity have been found and species B in electric eel extracts (obtained after trypsic treatment) has been identified with the molecular form of the enzyme as recently purified and cristallized.
Abstract: After extraction of the torpedo electric organ, three main molecular species, A, C, D, carrying the acetylcholinesterase activity have been found. This is observed in fresh-frozen organ extracts, toluene-autolysed organ extracts, or autolysis exsudate. A fourth species, B, appears during autolysis in the exsudate, or following treatment of the extracts by proteolytic enzymes, trypsin or α-chymotrypsin. A series of agents such as pH, p-hydroxymercuribenzoate, or pharmacological drugs, have been tried in order to modify the distribution of activity among the different species; all have failed, demonstrating the stability of these species.
The two species which migrate faster in a sucrose gradient, C and D, show the phenomenon of aggregation in low salt concentration (0.01 M), whereas and A and B, the slower species, do not.
The buoyant density of the active species in our extract is the same as that of the purified enzyme; a high content of lipids seems therefore excluded.
Species B in electric eel extracts (obtained after trypsic treatment) has been identified with the molecular form of the enzyme as recently purified and cristallized.
This raises the question of the biological significance of this preparation, especially since one of the original species, A, has a smaller mass and is also converted into B during proteolysis.
180 citations
TL;DR: Preparations of bovine pancreatic deoxyribonuclease obtained by the ammonium sulfate precipitation procedure of Kunitz can be further purified by chromatography on sulfoethyl-Sephadex at pH 4.70 and show that the enzyme contains glucosamine and mannose and establish DNase as a glycoprotein.
165 citations
TL;DR: Biological activity of thyrotropin-releasing hormone was abolished by diazotized sulfanilic acid, N-bromosuccinimide, or acid hydrolysis, but was not affected by periodate or by incubation with proteolytic enzymes.
155 citations
TL;DR: The striking distributional regularities of the basic and hydrophilic residues in the sequence and the importance of the positions of half cystinyl residues which form the disulfide bonds for maintaining the protein in its active configuration, are considered in connection with the structure-activity relationship of the protein.
TL;DR: It was concluded that only the outside surface of the vesicles is susceptible to the proteolytic action and that cytochrome b 5 and the NADPH-specific flavoprotein are located in the susceptible area.
Abstract: Digestion of rabbit liver microsomal smooth vesicles with Bacillus subtilis protease released proteins and peptide fragments from the vesicles, without solubilizing phospholipids and cholesterol. The proteolysis was, however, limited when about 30% of the protein had been solubilized. The same limitation was observed when the vesicles were treated with trypsin, chymotrypsin, or their combinations with the bacterial protease. The limited proteolysis was accompanied by selective solubilization of cytochrome b5 and microsomal NADPH-specific flavoprotein, leaving the CO-binding hemoprotein and some other enzymes still attached to the vesicular membranes. Sucrose density gradient centrifugation of protease-treated vesicles indicated that all the vesicles had been attacked by the protease to similar extents. The behavior of intact and digested vesicles in dextran density gradient centrifugation suggested that the vesicles, even after proteolytic digestion, existed in the form of closed sacs which were impermeable to macromolecules such as dextran and proteases. It was concluded that only the outside surface of the vesicles is susceptible to the proteolytic action and that cytochrome b5 and the NADPH-specific flavoprotein are located in the susceptible area.
TL;DR: In conclusion, inhibition of bovine trypsin by naturally occurring inhibitors is no index of their activities against human tryps in terms of the activity of the Kunitz soybean inhibitor.
TL;DR: It is suggested that ochratoxin A is hydrolyzed in vitro by carboxypeptidase A and a-chymotrypsin and possibly cathepsin C from lyosomes.
TL;DR: The amyloid fibrils in their native state proved to be remarkably resistant to digestion by a number of proteolytic enzymes, and peptide maps demonstrated differences among amyloids preparations from different subjects.
Abstract: Amyloid fibrils were isolated from the tissues of nine patients with amyloidosis in a state of high purity by homogenization of the tissue followed by extraction with distilled water. Physical, chemical, and ultrastructural studies suggest that amyloid fibrils from different individuals resemble each other, but are not identical. In tissue sections as well as by negative staining of isolated fibrils, morphologic variations were observed. Among the isolated fibrils at least three types were noted. The majority resembled those described previously. However, one subject had two types of fibrils which differed in size and appearance.
Most of the preparations sedimented as a single component with a sedimentation coefficient of 45–50S or as a larger polymer. However, two of the preparations had sedimentation coefficients of 8–9S, and a third one had a major 95S component and a minor 9S fraction.
While the preparations of amyloid were not sufficiently pure for amino acid analyses, peptide maps demonstrated differences among amyloid preparations from different subjects.
The amyloid fibrils in their native state proved to be remarkably resistant to digestion by a number of proteolytic enzymes. Several chemical methods were tried to produce smaller subunits. Of these, the most successful one was the use of 0.1 M NaOH which yielded a smaller, soluble fraction with sedimentation coefficients ranging from 1.1 to 2.8S. Accompanying this degradation, there was little loss of peptides or carbohydrates. Based on the results of the chemical analyses, it is estimated that the subunit produced by sodium hydroxide had a molecular weight of approximately 35,000–40,000.
TL;DR: Bacteriocin activity was readily demonstrated in agar agarose or starch plate assays, but activity could not be demonstrated in broth cultures, indicating that the bacteriocins were extremely labile in liquid media, but they were found to be stabilized by low concentrations of agar, starch, glycerol or dextran.
TL;DR: Results suggest that a site (or sites) on the cell surface containing peptide components is necessary for the action of insulin on glucose transport and lipolysis, and is probably distinct from the elements involved directly in glucose transport, lipolytic activity, and turnover of cyclic AMP.
TL;DR: A decreased inhibition of pancreatic elastase has been detected in the serums of six patients with α1-antitrypsin deficiency, extending the defect of inhibition by serum to a second, biologically active proteolytic enzyme in this form of familial emphysema.
Abstract: A decreased inhibition of pancreatic elastase has been detected in the serums of six patients with α1-antitrypsin deficiency. Five have severe clinical and physiological pulmonary emphysema. This observation extends the defect of inhibition by serum to a second, biologically active proteolytic enzyme in this form of familial emphysema.
TL;DR: It has been shown that the ratio of solid to liquid in the reaction mixture can be kept constant to give accurate values when pipetting aliquots from a solid-liquid mixture and that Azocoll is more sensitive to trypsin than previously reported.
TL;DR: Gel filtration chromatography indicates matrone activity can be separated into at least two components, both macro-molecules, and at least one of the molecular components which make up active matrone is a protein.
TL;DR: Halobacterium strains produce a truly extracellular proteinase which degrades gelatine and casein and depends upon divalent cations and a high concentration of NaCl or KCl for activity and stability.
Abstract: SUMMARY: Halobacterium strains produce a truly extracellular proteinase which degrades gelatine and casein. It has a pH optimum of about 8 and depends upon divalent cations and a high concentration of NaCl or KCl for activity and stability. Proteolytic enzymes were also found in cell homogenates obtained by ultra sonic treatment. A caseinolytic probably different from the extracellular one, is associated with particles which sediment upon ultracentrifugation. A soluble peptidase of weight is also present in the extract. Both enzymes are dependent upon divalent cations and a high concentration of NaCl or KCl for activity. In contrast to other halophilic enzymes, the proteolytic enzymes of Halobacterium salinarium are more active in the presence of NaCl than KCl at equilmolar concentrations.
TL;DR: The basic principles of the topochemical approach to the investigation of the structure-function relation in peptide systems are formulated and the fruitfulness of using such compounds as tools in elucidating the physicochemical basis of functioning of biological membranes is shown.
Abstract: The basic principles of the topochemical approach to the investigation of the structure-function relation in peptide systems are formulated. This approach makes use of the new possibility of transforming natural peptides, consisting in the modification of the molecule as a whole and utilization of the resultant analogs to elucidate the boundaries of the stereoelectronic complementarity of the biologically active peptide to the corresponding receptor. In particular, on the example of depsipeptide antibiotics and their topochemical analogs the fruitfulness of using such compounds as tools in elucidating the physicochemical basis of functioning of biological membranes is shown. The topochemical principle has also been applied in preparing specific competitive inhibitors of proteolytic enzymes, whose study may shed light on the nature of the forces binding the substrate to the contact site of the corresponding enzyme.
TL;DR: It is inferred that proteases play an important role in early myelin breakdown and may also be implicated in the pathogenesis of multiple sclerosis plaques by digesting basic protein, by releasing lipid from its attachment to such protein, and by liberating an active encephalitogenic peptide.
Abstract: Proteins are important constituents of the myelin sheath and serve to maintain its structural integrity. One of the protein components is susceptible to tryptic digestion and may be regarded as a particularly vulnerable part of the myelin sheath. The initial events in myelin breakdown may involve disruption of lipid-protein attachments followed later by chemical degradation of released lipids. In Wallerian degeneration the activity of proteolytic enzymes increases by 12 hr after nerve section. Proteolytic enzyme activity increases in the prodromal phase of diphtheritic neuropathy. Extracts of degenerating nerve cause proteolysis of normal myelin with loss of trypanophilic basic protein and lipid; selective loss of basic protein occurs very early in Wallerian degeneration and has also been found in and around plaques of multiple sclerosis. Proteolytic activity is increased at the edges of ‘active’ multiple sclerosis lesions. It has been shown that the basic encephalitogenic protein is susceptible to digestion by neural proteases, yielding an active encephalitogenic fragment. It is inferred from these collective observations that proteases play an important role in early myelin breakdown and may also be implicated in the pathogenesis of multiple sclerosis plaques by digesting basic protein, by releasing lipid from its attachment to such protein, and by liberating an active encephalitogenic peptide. The factors responsible for the activation and release of proteases remain unknown.
TL;DR: Radioactivity in rat liver was investigated 20 min after intraperitoneal injection of 20 and 200 μC of [3H]cortisol and was found to be weakly associated with a macromolecular fraction in a form resistant to nucleases but sensitive to pronase, trypsin and neuraminidase.
TL;DR: Electrophoresis experiments suggested preferential pronase attack on some higher molecular weight membrane proteins; these same electrophoretic components were preferentially released at very low concentrations of sodium dodecyl sulfate,, indicating differences in the organization of proteins within the membrane.
TL;DR: A trypsin inhibitor present in ungerminated barley in relatively high concentration has been obtained in apparently pure form and is totally inactive against chymotrypsin and pepsin, proteolytic enzymes of germinating barley, and a number of microbial proteinases.
Abstract: A trypsin inhibitor present in ungerminated barley in relatively high concentration (0.45 mg/g) has been obtained in apparently pure form. The inhibitor is a heat-stable protein. Molecular weights of 14400 and 14055 were calculated from the sedimentation equilibrium and amino acid composition respectively. 1 μg of purified inhibitor inhibits 1.6–1.9 μg of pure trypsin in hydrolyses of casein, haemoglobin, and benzoyl-dl-arginine-p-nitroanilide. The inhibitor preparations are totally inactive against chymotrypsin and pepsin, proteolytic enzymes of germinating barley, and a number of microbial proteinases.
TL;DR: Support is found for the hypothesis that proteinaceous foods stimulate protease synthesis rather than activate a precursor and a powerful inhibitor for the cockroach protease in found in the anterior midgut and the caeca.
TL;DR: The red cells from a 23‐year‐old pregnant woman gave very unusual blood grouping reactions and her serum contained a ‘new’ antibody active against a very high incidence antigen.
Abstract: Summary. The red cells from a 23-year-old pregnant woman gave very unusual blood grouping reactions and her serum contained a ‘new’ antibody active against a very high incidence antigen.
The reactions of various unrelated blood group antigens are modified, in some cases enhanced and in others depressed, the total picture being strongly reminiscent of the effects of proteolytic enzyme treatment. It is suggested that these effects can only be due to some factor affecting the red cell structure possibly by modifying the cell envelope.
This modifying factor is genetically determined and shows dosage effect. Of nine siblings three are apparently homozygous and four heterozygous for this factor, the other two being ‘normal’. It is extremely rare, no further examples having been found in 18,472 blood donors.
The antibody is undoubtedly immune, probably due to a blood transfusion two years previously. 12,509 donors have been tested with it but the only compatible red cells are those of the three homozygous siblings, i.e. the propositus, one sister and one brother. The antibody is therefore related to the modifying factor.
Two codominant genes Ena and Enb are postulated, Enb being exceedingly rare. It is suggested that these genes determine not only the presence of the corresponding antigens Ena and Enb but also the structure of the red cell.
TL;DR: Pleitropic interactions among genes controlling the formation of bacterial spores and of sporulation-associated products are studied in this paper, where the results confirm the existence of a unidirectional pleiotropic system.
Abstract: Pleitropic interactions among genes controlling the formation of bacterial spores and of sporulation-associated products are studied. In order to obtain sporulation mutants, spores have been germinated in the presence of chloramphenicol and then treated with nitrosoguanidine. In the most favorable conditions 25% of sporulation mutants have been found among the 40% surviving bacteria. This number is at least four times higher than the number of auxotrophic mutants, therefore a rough estimate of the number of genes involved in sporulation is 800. Rapid plate-tests have been developed for the oxidation of terrazolium salts, the formation of various proteolytic enzymes and the production of antibiotics. Although the exact biochemical nature of the products is not yet known, the results suggest that distinct factors, probably various enzymes (including several proteases) are detected by these tests. All of them are associated with spore formation and absent from a large number of sporulation mutants. Using these tests, the phenotypes of 500 randomly selected sporulation mutants were determined. No important differences were found between asporogenous and oligosporogenous mutants. The number of mutants deficient for several sporulation-associated characters is large, pleiotropic interactions following a defined pattern are observed. Statistical analysis indicates the existence of a unidirectional pleiotropic system. All the results agree with the hypothesis of sequential gene activation. Consequently, the sporulation-associated characters can be ordered into a linear sequence, presumably reflecting the consecutive steps in spore formation. The order obtained is the following: gelatinase, proteases acting on casein and on denatured albumin, oxidation of tetrazolium No 7, digestion of protamine, production of antibiotics (against a Staphylococcus and a Bacillus), hydrolysis of hemoglobin, oxidation of tetrazolium No 2, digestion of native albumin, synthesis of elastase. Another category of mutants, blocked in a late step of sporulation and apparently derepressed for the formation of elastase, is also described. In conclusion, arguments are put forward in favor of sequential gene activation. Sporulation genes, related by unidirectional pleiotropic interactions, form a sporulon. Generalization of this concept to other differentiating systems (a differon), its predictions and possible experimental confirmation are considered.