Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This paper presents the partial characterization and the identification of an 80-kDa protein detected in bull spermatozoa using a monoclonal antibody directed against a 16-amino acid long peptide from the N-terminal domain of the protooncogene p60(src) from the Rous Sarcoma Virus When subjected to two-dimensional electrophoresis, this 80-kDa protein migrated as several isoforms, with an isoelectric point ranging from 7.4 to 8.2. Amino acid sequence analysis of a peptide obtained following trypsin digestion of the bull sperm protein showed homology to the PH-20/hyaluronidase precursor sperm protein. As for PH-20, this bull sperm 80-kDa protein is located at the plasma membrane surface in the postacrosomal region of the head. An increased immunolabeling in the anterior head region of fixed/permeabilized spermatozoa was observed when these cells were incubated under capacitating conditions, whereas most sperm cells challenged with the calcium ionophore A23187 to acrosome react lost their labeling almost completely. As for the PH-20 protein, the 80-kDa bull sperm protein possesses a hyaluronidase activity that is higher at pH 7.0 than at pH 4.0 in an in-gel assay. Unlike what has been observed in the guinea pig, mouse, and human PH-20, this 80-kDa protein was not released from the surface of bull spermatozoa by treatment with phosphatidylinositol-specific phospholipase C or with trypsin. However, this protein was not sedimented by a 100,000 x g centrifugation after nitrogen cavitation, which suggests that the 80-kDa protein is loosely attached to the sperm membrane by a yet-unknown mechanism. These results suggest that the 80-kDa bull sperm protein shares many homologies with the sperm PH-20 protein reported in the literature and, most likely, is the bull sperm homologue of the PH-20.
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PMID:Characterization of an 80-kilodalton bull sperm protein identified as PH-20. 1146 35

In search for Xenopus laevis hyaluronidase genes, a cDNA encoding a putative PH-20-like enzyme was isolated. In the adult frog, this mRNA was only found to be expressed in the kidney and therefore named XKH1. When expressed by means of cRNA injection into frog oocytes, XKH1 solely exhibited at physiologic ionic strength hyaluronidase activity at neutral pH and in weakly acidic solutions. The enzyme was inactive below pH 5.4. In addition to hyaluronic acid hydrolysis, chondroitin sulfate also was degraded at low yield as assessed by fluorophore-assisted carbohydrate electrophoresis analysis of the degradation products. The enzyme is sorted to the outer surface of the cell membrane of XKH1 expressing oocytes. From there, it could not be removed by phospholipase C nor was secreted hyaluronidase activity detectable. We conclude that XKH1 represents a membrane-bound hyaluronan-degrading enzyme exclusively expressed in cells of the adult frog kidney where it either may be involved in the reorganization of the extracellular architecture or in supporting physiological demands for proper renal functions.
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PMID:Xenopus kidney hyaluronidase-1 (XKH1), a novel type of membrane-bound hyaluronidase solely degrades hyaluronan at neutral pH. 1156 78

PH-20 is a glycoprotein located on the surface of the sperm plasma membrane and on the inner acrosomal membrane. The best understood function of sperm surface PH-20 is its hyaluronidase activity, which results in hydrolysis of the hyaluronic acid-rich cumulus matrix during sperm penetration of this extracellular oocyte investment. In this study, we investigated whether alterations in the secondary and tertiary structures of sperm surface PH-20 would affect its enzyme activity. Proteins were isolated from the sperm plasma membrane by treatment of living cells with phosphatidylinositol-specific phospholipase C (PI-PLC). PH-20 was purified from the PI-PLC released proteins by immunoaffinity chromatography. Two-dimensional electrophoresis of purified PH-20 revealed 6 isoforms with isoelectric points ranging from 5.1 to 6.0. Removal of the N-linked glycans from PH-20 with N-glycosidase F shifted the molecular weight from 64 kd to approximately 54 kd, its deduced molecular weight based on sequence analysis, suggesting that most if not all, of the potential N-glycosylation sites are linked to oligosaccharides. The lectins Con A and PSA recognized purified sperm surface PH-20 after Western blotting, suggesting that mannose is a major sugar within or at the terminal end of the linked glycan. The lectins UEA and LPA did not recognize PH-20 Western blot, suggesting that fucose and sialic acid are not terminal sugars of sperm surface PH-20. Deglycosylation of sperm surface PH-20 resulted in a complete loss of its hyaluronidase activity. The reduction of disulfide bonds with beta-mercaptoethanol or dithiothreitol also resulted in loss of enzyme activity. We conclude that the hyaluronidase activity of sperm surface PH-20 is dependent on structural features established by sulfhydryl linkages, as well as glycosylation.
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PMID:Importance of glycosylation and disulfide bonds in hyaluronidase activity of macaque sperm surface PH-20. 1186 14

Previously we demonstrated that the murine sperm adhesion molecule 1 (Spam1 or PH-20) is synthesized by the epididymal epithelium, preferentially in the distal region, and is released into the luminal fluid. We also showed that whereas testicular and epididymal Spam1 have hyaluronidase activity at neutral pH, they are under different transcriptional regulation. The aim of this study was to further compare characteristics of the two forms of this glycosyl-phosphatidylinositol-linked protein and their transcripts, and to determine whether secreted epididymal Spam1 is released with its lipid anchor. With GeneRacer amplification of the 3' end of the complementary DNA we show that the poly(A) tails are significantly (P <.05) shorter in the epididymis than in the testis. Two-dimensional polyacrylamide gel electrophoresis with immunoblotting reveals one to three isoforms for epididymal Spam1 with the isoelectric point (pl) ranging from 7.3 to 9.0, and four isoforms ranging from 6.6 to 9.0 pl for testicular Spam1. Two isoforms with a pl ranging from 7.6 to 9.0 were observed for caudal sperm. Lectin blotting analysis shows that Phaseolus vulgaris erythroagglutinin, Lycopersicon esculentum lectin (LEL), and Solanum tuberosum lectin, which all bind to N-linked chains, recognize a 67 kd band in the epididymis and caudal sperm, but not in the testis. Treatment of the protein extracts with anti-Spam1 serum prior to blotting with LEL led to the disappearance of the banding, indicating Spam1 specificity of the staining. The lectin peanut agglutinin, which preferentially binds to O-linked side chains, recognizes a 67 kd band in all three cell types. Enzymatic deglycosylation studies confirmed the presence of an O-linked glycan in all three cell types. Ultracentrifugation of the luminal fluid reveals that epididymal Spam1 is secreted predominantly as insoluble particles, which when treated with phosphatidylinositol-specific phospholipase C or Triton X-100, reveal that the majority of epididymal Spam1 is released with its lipid anchor, a form in which it can bind to sperm.
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PMID:Mouse epididymal Spam1 (pH-20) is released in the luminal fluid with its lipid anchor. 1251 83