Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The aryl 4-guanidinobenzoate, 4'-nitrophenyl 4-guanidinobenzoate (NPGB), is a potent inhibitor of sperm acrosin, an enzyme with an essential function in the fertilization process. NPGB prevents fertilization in a number of animal species and is a good lead compound for the development of contraceptive agents. In order to assess the efficacy of other aryl 4-guanidinobenzoates as acrosin inhibitors, 24 of these compounds were synthesized. Their inhibitory activity toward human acrosin was determined and compared with their activity toward human pancreatic trypsin in order to assess whether inhibitor sensitivity differed between these similar enzymes. Nine of the inhibitors were synthesized from phenols approved by the FDA for therapeutic use. The acute toxicity of these inhibitors in mice was determined and compared to that of nonoxynol-9, the most commonly used active ingredient in today's vaginal contraceptive preparations. All of the compounds proved to be potent inhibitors of human acrosin although 3 orders of magnitude difference were observed between the most and least effective inhibitors. Little specificity was present in regard to their inhibition of acrosin and trypsin. All the aryl 4-guanidinobenzoates synthesized from FDA-approved phenols were less toxic than nonoxynol-9, and it is concluded that these 4-guanidinobenzoates are of interest for further development and testing as nonhormonal contraceptive agents.
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PMID:Synthesis and inhibition of human acrosin and trypsin and acute toxicity of aryl 4-guanidinobenzoates. 351 12

The presence of hydrolytic enzymes in and associated with the sperm head has long argued for their functioning in fertilization. Several observations led investigators to propose that the acrosomal trypsin-like enzyme, acrosin in mammals, functioned in fertilization in aiding the sperm to penetrate the zona pellucida. While many have raised significant objections to this role, the action of acrosin on its presumed physiological substrate has not been characterized in a biochemical fashion. The intent of this study was to examine the effect of sperm proteases on the innermost egg envelopes in a parallel study, with the pig, Sous scrofa and the South African clawed frog, Xenopus laevis. With the pig, a great deal of information exists concerning the boar enzyme, acrosin, but little is known about the chemical structure of the zona pellucida. The opposite situation exists in X. laevis where the vitelline envelope is well characterized chemically, but little is known about the putative sperm lysins.
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PMID:The action of acrosin on the zona pellucida. 354 37

Rabbit antiserum against trypsin-kallikrein inhibitor (TKI) was prepared. Purified immunoglobulin G (IgG) fraction was bound to Sepharose 4B. An antigen immunologically related to TKI was obtained from porcine blood plasma by adsorbing it onto the immunosorbent column. Its immunoreactivity with TKI antibodies was confirmed by immunoelectrophoresis. The antigen was an inhibitor of trypsin and acrosin.
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PMID:Detection of antigen immunologically related to basic pancreatic inhibitor (Kunitz) in porcine blood plasma by immunoaffinity chromatography. 371 Nov 98

A proteinase inhibitor was isolated from extracts of the leech Hirudo medicinalis by gel filtration and anion exchange chromatography. This inhibitor is similar to the bdellins in that it blocks the activity of trypsin, plasmin and sperm acrosin but has a molecular mass, as estimated by SDS polyacrylamide electrophoresis, of about 20 kDa, whereas the bdellins have molecular masses in the range 5-6 kDa. It is therefore designated as high-molecular mass bdellin B-3 (HMB). The amino-acid sequence of the inhibitor was elucidated as far as position 56. This revealed that the molecule consists of a bdellin B-3 moiety, corresponding to the N-terminal 46 residues, which is then extended at the C-terminus by a polypeptide chain of the composition Asx15, Glx25, Gly6, Val, His26-27 and Lys4. It has been formerly concluded from a partial amino-acid sequence that bdellin B-3 is a Kazal-type inhibitor. However, the complete sequence of bdellin B-3, represented by the N-terminal 46 residues of HMB, discloses that bdellin B-3 is a non-classical Kazal-type inhibitor when the number of amino-acid residues between half-cystines are considered. Presuming that formation of disulfide bridges principally follows the same pattern as in classical Kazal-type inhibitors the bdellin B-3 molecule was modeled based on the known three-dimensional structure of the third ovomucoid domains. This showed that a compact arrangement of the peptide chain of bdellin B-3 is conceivable.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The primary structure of bdellin B-3 from the leech Hirudo medicinalis. Bdellin B-3 is a compact proteinase inhibitor of a "non-classical" Kazal type. It is present in the leech in a high molecular mass form. 382 73

We previously reported the activation of adenylate cyclases from rat brain (Johnson, R. A., Awad, J. A., Jakobs, K. H., and Schultz, G., (1983) FEBS Lett. 152, 11-16) and from human platelets (Jakobs, K. H., Johnson, R. A., and Schultz, G. (1983) Biochim. Biophys. Acta 756, 369-375) by a factor derived from bovine sperm. In this report we describe the conditions for the extraction of the factor from bovine sperm and characteristics of its effects on adenylate cyclase which are consistent with its being a protease. The activating capacity of sperm particles was extracted from previously washed and frozen sperm into a 30,000 X g supernatant fraction by various salts, but not by the nonionic detergent Lubrol-PX. The amount of extracted factor: (a) was greatest with NH4HCO3 greater than NaCl greater than Na acetate; (b) was optimal with 0.5 M salt; (c) was not appreciably affected by the pH of the extraction buffer between pH 5.0 and 8.5; and (d) exhibited the greatest specific activity at the lower pH. The extracted sperm factor could be concentrated without loss by ultrafiltration on Amicon PM-10 membranes. The effect on adenylate cyclase of concentrated and desalted sperm extracted was inhibited 50% by various salts at 10 to 30 mM. The effects of the sperm factor to activate platelet adenylate cyclase, to block its inhibition via the alpha-adrenoceptor, and to block inhibition of stimulated forms of the enzyme by stable guanine nucleotides were prevented by protease inhibitors. A 50% reduction in the sperm factor's activation of platelet adenylate cyclase was caused by 30 nM soybean trypsin inhibitor, 30 nM alpha 2-macroglobulin, 300 nM leupeptin, 1 microM antipain, 15 microM aprotinin, and 100 microM benzamidine. Up to 3 mM phenylmethanesulfonyl fluoride was without effect on activation of the platelet cyclase by the sperm factor. The effects of the sperm factor persisted after its removal by the washing of pretreated platelet membranes and after its inactivation by the subsequent addition of leupeptin. The data strongly support the conclusion that the bovine sperm factor is a trypsin-like protease. alpha-Chymotrypsin, trypsin, and sperm acrosin were comparably effective in stimulating the platelet adenylate cyclase 5- to 8-fold, with concentrations eliciting maximal stimulation being: 200 ng trypsin/ml; 2 micrograms alpha-chymotrypsin/ml; and 2 micrograms acrosin/ml.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Extraction of the adenylate cyclase-activating factor of bovine sperm and its identification as a trypsin-like protease. 388 Jul 36

The proteolytic action of boar sperm acrosin on its natural substrate, the zona pellucida, was investigated. Acrosin exhibited substrate specificity for the zona pellucida and differentially hydrolyzed the glycoprotein families composing the zona pellucida. In contrast to acrosin, trypsin was a less-specific protease in terms of zona pellucida hydrolysis.
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PMID:Limited and specific proteolysis of the zona pellucida by acrosin. 388 82

The purpose of this study was to examine how trypsin inhibitors affect the guinea pig sperm acrosome reaction in vitro. Using spermatozoa pretreated with lysophosphatidyl choline, we found that both naturally occurring high molecular weight and the smaller synthetic trypsin inhibitor p-aminobenzamidine (PAB) delayed the onset of the acrosome reaction as monitored by light microscopy. Examination with electron microscopy revealed that acrosomal matrix dispersal rather than membrane fusion was affected. Despite the morphologic delay in acrosomal content release, PAB unexpectedly permitted 96% of soluble acrosomal antigen to be released into the supernatant. In addition, total acrosin release in the presence of PAB was 74% of control, with the vast majority as latent rather than active enzyme. A morphologically intact but membrane-free target of acrosomal matrix (AM), which is sensitive to trypsin inhibitor, was partially purified using Triton-x-100 at pH 5.2. AM remained morphologically stable at pH 5.2; however, shift up to pH 7 resulted in rapid dissolution within several minutes as monitored by light and electron microscopy and light scattering. Trypsin inhibitor prevented dispersion of AM at pH 7. The results suggest that, during the acrosome reaction, one distinct region of the acrosomal contents disperses after membrane vesiculation in a pH and trypsin inhibitor-insensitive fashion while a pH sensitive trypsin-like activity (acrosin?) disperses another discrete region of acrosomal matrix.
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PMID:pH and protease control of acrosomal content stasis and release during the guinea pig sperm acrosome reaction. 388 29

The morphologic and biochemical effects on the structure and constituent glycoproteins of the zona pellucida (ZP) by a specific sperm enzyme, acrosin, and a nonsperm enzyme, trypsin, have been evaluated. Intact porcine ZP matricies, exposed to either acrosin or trypsin, were analyzed microscopically. Changes in specific glycoproteins were monitored by high-resolution two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and the silver-based color stain, GELCODE. Although these enzymes did not alter the macroscopic properties of the ZP matrix, the 2D-PAGE ZP protein patterns were markedly altered. The high molecular weight glycoprotein families (II and III) were sensitive to proteolytic digestion, whereas the major glycoprotein family (I) of the porcine zona was only partially proteolyzed by acrosin and trypsin. Furthermore, it was demonstrated that acrosin had unique substrate specificity compared to that of trypsin, since the ZP peptide patterns were found to be different. These studies are the first to demonstrate which integral glycoproteins of the native porcine ZP matrix are specifically proteolyzed by acrosin from the homologous species and that this proteolysis occurs without the dissolution of the native porcine matrix.
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PMID:Proteolysis of specific porcine zona pellucida glycoproteins by boar acrosin. 388 99

Acrosin (acrosomal proteinase; EC 3.4.21.10) is a sperm-specific serine proteinase implicated in sperm penetration of the mammalian oocyte. Previously, we had shown that human acrosin, unlike human trypsin (EC 3.4.21.4), was inhibited by beta-D-fructose and related carbohydrates. The present study was undertaken to more fully elucidate the mechanism of action of fructose as an acrosin inhibitor, and to further differentiate the kinetic properties of acrosin from those of trypsin. Fructose produced a complex pattern of inhibition. At relatively low concentrations (10-60 mM), fructose acted as a competitive inhibitor with an apparent inhibition constant of 13 mM. In contrast, at high concentrations (80-320 mM), fructose behaved as a noncompetitive inhibitor, with an apparent inhibition constant of 205 mM. A Hill plot of enzyme activity as a function of fructose concentration suggested only a single binding site for fructose (slope = -0.90). The pattern of inhibition is not consistent with an enzyme containing only a single catalytic site, based either upon steady-state or rapid equilibrium assumptions; however, good agreement between observed and simulated data were obtained based upon the assumption of two catalytic sites with equal or similar binding and catalytic constants. The data suggested that fructose interacts with a single binding site (Ki = 8 mM) which alters both catalytic sites to produce an enzyme species having a higher apparent Michaelis constant and lower kcat as compared to the uninhibited enzyme. Fructose had no effect upon the rate of acrosin inactivation by either diisopropylfluorophosphate or tosyl-lysine-chloromethylketone, suggesting that neither substrate binding nor acylation were altered by this agent. The above data indicate substantial differences between the catalytic properties of human acrosin and those of trypsin.
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PMID:Evidence for multiple catalytic sites of human acrosin from kinetic evaluation of fructose-induced acrosin inhibition. 389 12

A new anionic acrosin inhibitor was found in an acidic extract of boar spermatozoa. The protein was purified by hydrophobic and ion exchange chromatography. According to gel filtration and SDS-electrophoresis the inhibitor preparation shows a molecular mass of Mr approximately 6000-7000 daltons. The isoelectric point is close to pH 4.5. It is an effective inhibitor of boar acrosin and bovine trypsin, but it does not inhibit porcine plasmin and pancreatic kallikrein or bovine chymotrypsin. An inhibitor with identical properties was found in high concentration (97% of the total acrosin inhibiting activity) in the fluid of cauda epididymis and also as a minor acrosin inhibiting component (2% of total acrosin inhibiting activity) in seminal plasma. The results indicate that binding of the inhibitor to spermatozoa may have taken place in the epididymis.
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PMID:Demonstration of an anionic acrosin inhibitor in spermatozoa epididymal fluid and seminal plasma of the boar. 390 26


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