Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Approx. 90% of the highly motile goat cauda-epididymal spermatozoa lose motility completely due to removal of the surrounding epididymal plasma (EP) by washing with a modified Ringer's solution. Motility can however, be reconstituted in approx. 80% of these washed cells that lost motility during washing, with the addition of goat cauda-EP. The activity of the EP-factor(s) that cause reconstitution of motility in the washed immotile spermatozoa, is nondialysable and sensitive to the action of trypsin. The activity of the factor(s) is not dependent on exogenous Ca2+ (1 mM), Mg2+ (1.2 mM), cyclic AMP (5 mM) or dibutyryl cyclic AMP (5 mM). The novel system may thus serve as an excellent analytical tool to yield an insight into the molecular basis of the regulation of spermatozoal motility.
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PMID:Evidence for the reconstitution of motility by epididymal plasma-protein factor(s) in immotile washed spermatozoa from goat cauda epididymis. 665 Aug 87

Aminopeptidase activity was demonstrated in the spermatozoa of the sea urchin, Strongylocentrotus intermedius. The enzyme solubilized from sperm cells was inactivated with bestatin, amastatin, p-chloromercuribenzenesulfonate, ZnCl2, and trypsin, whereas that in the intact cells was scarcely affected by these agents. On the other hand, bestatin ethyl ester inactivated the two enzyme forms to almost the same extent. It also inhibited the fertilization process in this species. These results suggest that the aminopeptidase associated with the spermatozoa is shielded with a permeable barrier and plays some role in fertilization.
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PMID:Sea urchin sperm aminopeptidase: comparative studies of sperm-associated and -solubilized enzymes. 667 49

A new proteinase inhibitor (Mr 7500) was isolated to apparent homogeneity from boar spermatozoa by repeated gel filtration on Sephadex G-50 and affinity chromatography on concanavalin-A--Sepharose 4B. The inhibitor strongly inhibits boar acrosin in a competitive 1:1 stoichiometric reaction with Kass = 7 x 10(10)1 mol-1. The inhibitor is a glycoprotein and represents a first member of a new class of proteinase inhibitor with a rather short polypeptide backbone of only 42 amino acid residues and a low cystine content. The basic protein (isoelectric point 9.4) contains a single disulfide loop, which is easily reducible by sodium borohydride. Upon reduction the inhibitory activity is lost, but rapidly regained after air reoxidation of the corresponding half-cystine residues. The reactive site residue was established to be arginine by inhibition with 2,3-butanedione. The inhibitor is rather specific for acrosin and inhibits bovine trypsin only to a limited extent. However, incubation with catalytic amounts of trypsin (or acrosin) at acid pH (pH2-3) rapidly leads to a limited proteolysis at the reactive site with formation of 67% modified (reactive site hydrolysed), but still active inhibitor. The equilibrium constant was established to be Khyd = 2.0.
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PMID:A new acrosin inhibitor from boar spermatozoa. 675 19

A proacrosin conversion inhibitor present in boar spermatozoa has been purified and initially characterized. Purification methods included sequential acid extractions of washed spermatozoa at pH 4.0, pH 3.5, and pH 2.5 followed by successive gel filtrations of the pH 2.5 sperm extract supernatant over Sephadex G-75 and G-50. The resulting 8.8-fold purified materials were judged to be homogeneous by sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis, had an estimated molecular weight of 12,800, and a constant specific activity of 65 units/mg. Treatment with the proteinases acrosin, trypsin, or chymotrypsin destroyed the highly purified proacrosin conversion inhibitor, indicating that it is a protein. Additional properties of the inhibitor included stability to long periods of storage at pH 3.0 and 4 degrees C, stability to boiling and lyophilization, and an absolute requirement for divalent cations to maintain activity. The highly purified proacrosin conversion inhibitor does not inhibit acrosin. Therefore, it apparently acts to prevent proacrosin conversion by selectively inhibiting the zymogen's self-catalyzed conversion mechanism.
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PMID:Proacrosin conversion inhibitor. Purification and initial characterization of a boar sperm protein which prevents the conversion of proacrosin into acrosin. 680 Oct 40

A high-molecular-weight form of acrosin (alpha-acrosin, EC 3.4.21.10) was extracted from spermatozoa obtained from frozen semen and purified over 300-fold. Purification was effected by sequential use of Sephadex G-150, CM-cellulose and DEAE-cellulose chromatography. Properties of human acrosin were compared with those of human pancreatic trypsin. The molecular weight (Mr) of acrosin (70000) was greater than that of trypsin (Mr 21000). Isoelectric points for acrosin (pI = 9.0) and trypsin (pI = 8.2) were also different. alpha-N-Benzoyl-L-arginine ethyl ester was hydrolysed 50% more rapidly by acrosin than by trypsin. Acrosin had similar kcat. values for the hydrolysis of esters with different acylating groups (i.e. benzoyl-L-arginine and p-tosyl-L-arginine esters). In contrast, trypsin had dissimilar kcat. values for the hydrolysis of esters with different acylating groups. Kinetic data argue against deacylation as the rate-limiting step in ester hydrolysis by acrosin. Acrosin was less sensitive than trypsin to inhibition by 7-amino-1-chloro-3-L-tosylamidoheptan-2-one ('TLCK'), di-isopropyl fluorophosphate and soya-bean trypsin inhibitor. D-Fructose and D-arabinose inhibited acrosin, but had no effect on trypsin. The data indicate that definite differences exist between human acrosin and trypsin.
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PMID:Characterization of a high-molecular-weight form of human acrosin. Comparison with human pancreatic trypsin. 680 60

Human and bovine spermatozoa have been collected and washed repeatedly with isotonic saline to remove seminal plasma inhibitors and activate the acrosin. Then the acrosin activity of the cells was assayed with alpha-N-Benzoyl-DL-Arg-beta-naphthylamide (BANA). It was found that the surface-bound enzyme was not inhibited by high molecular weight inhibitors of trypsin but was markedly inhibited by low molecular weight trypsin inhibitors. Divalent metals (Zn++, Cu++, Hg++, Co++, Cd++) were all efficient inhibitors of acrosin on the washed cells. It was shown that the removal of zinc or copper from acrosin completely restored activity. It is proposed that the different levels of zinc in the male and female genital tract regulate acrosin activity. Aged cells released a soluble acrosin which was inhibited by serum and seminal plasma inhibitors of trypsin-like enzymes as well as by zinc ions in an identical manner to the surface-bound enzyme.
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PMID:Inhibition of human and bovine sperm acrosin by divalent metal ions. Possible role of zinc as a regulator of acrosin activity. 681 4

Anionic sites of rat epididymal spermatozoa were measured at pH 7.4 using tritiated polycationized ferritin. The spermatozoa from the caput region had 1.25 +/- 0.06 X 10(6) anionic sites per cell and a binding constant of 1.26 +/- 0.01 X 10(6) M-1. Spermatozoa from the cauda region had 1.50 +/- 0.09 X 10(6) anionic sites per cell and a binding constant of 4.84 +/- 0.82 X 10(6) M-1. The values were mean +/- s.d. The anionic sites were partly sensitive to treatments with neuraminidase, trypsin and Triton X-100.
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PMID:Measurement of anionic sites of rat epididymal spermatozoa using tritiated polycationized ferritin. 688 40

Work from a number of laboratories has shown that fertilization is blocked in the presence of protease inhibitors, although the specific site of inhibition has not been identified. The present experiments were designed to discriminate between sperm binding to zonae pellucidae as opposed to sperm penetration through zonae, so as to assess the effect of protease inhibitors on these two distinct events. Exposure of capacitated mouse spermatozoa to a variety of protease inhibitors directed against trypsin blocked sperm binding to zonae in a concentration-dependent manner. A chymotrypsin-directed inhibitor was not capable of blocking sperm binding to zonae. The trypsin inhibitors did not affect sperm penetration though zonae nor gamete membrane fusion if the sperm had established a firm association with the zona surface before addition of the inhibitors. Previous incubation of zona-intact eggs with the inhibitors did not lead to a reduction in sperm binding, indicating that the activity affected by the inhibitors is borne by spermatozoa. Interaction between spermatozoa and the zona surface appeared to be the specific locus of inhibition; sperm binding to zona-free eggs (i.e., binding to the egg plasma membrane) was unaltered by the trypsin inhibitors. These results suggest a reevaluation of the function of proteases in fertilization focusing on their role in initial sperm contact with the zona pellucida.
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PMID:Involvement of trypsin-like activity in binding of mouse spermatozoa to zonae pellucidae. 694 26

A rabbit antibody to mouse 3T3 cell fibronectin was used in conjunction with a fluorescein-tagged second antibody to detect fibronectin-like activity on the surface of rabbit spermatozoa. Only ejaculated sperm displayed an intense and highly localized fluorescence over the acrosomal region. Cauda epididymal sperm of the rabbit as well as several other species did not exhibit any reaction. The fluorescent activity could be eliminated by trypsin treatment but was re-established by incubation in cell-free seminal fluid. Sperm recovered from females 10-12 h after mating showed a reduction or absence of antifibronectin fluorescence, suggesting that this component's loss could be a factor in sperm capacitation. Because fibronectins show strong binding to collagen, mixtures of ejaculated sperm and collagen were examined in the light and electron microscope. Living sperm appear to have a strong affinity for collagen and quickly adhere to the filaments by their heads, while continuing vigorous flagellations. Surface labeling of sperm with the galactose-oxidase-NaB[3H]4 technique, extraction with urea-detergent mixtures and affinity chromatography of extracts on gelatin-Sepharose revealed a single radioactive band of mot wt approximately 40,000 after SDS polyacrylamide gel electrophoresis and fluorography.
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PMID:A collagen-binding protein on the surface of ejaculated rabbit spermatozoa. 699 66

The presence of structures bridging the inner and outer acrosomal membranes of the equatorial segment of boar spermatozoa was clearly demonstrated in cells that have undergone a variety of treatment procedures to displace the electron-dense contents of the acrosome. En-face sections show bridges to be punctate and not linearly extensive as might be suggested by sections perpendicular to the flat plane of the head. About 4.5 x 10(5) bridges, each measuring 7 nm across and spaced 7 nm apart, are arrayed hexagonally in the equatorial segment, but bridges are not present within the principal segment of the acrosome. Short-term treatment with trypsin partially digests the bridges, but does not disrupt the spacing or strict parallel configuration of equatorial segment membranes. However, short-term treatment with pronase digests most bridges and effectively disrupts the typical configuration of the equatorial segment. Freeze-fracture of the cytoplasmic face of the acrosomal membranes of the equatorial segment reveals a pattern throughout the phospholipid layer of the membrane which is similar to the pattern of bridges present in en-face thin sections of the equatorial segments. The data suggest that numerous bridges link the inner and outer acrosomal membranes of the equatorial segment of the acrosome and they play a major, if not an exclusive, role in maintaining the close spacing and parallel arrangement of the membranes in this portion of the acrosome.
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PMID:On the presence of bridges linking the inner and outer acrosomal membranes of boar spermatozoa. 700 60


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