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:1.2.1.13 (glyceraldehyde-3-phosphate dehydrogenase)
6,511 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The current study adds to the growing body of evidence that RNA is present in mature ejaculated human spermatozoa. We report that a sodium dodecyl sulphate (SDS)/citric acid extraction method is superior to guanidinium isothiocyanate in terms of reproducibility of RNA recovery from motile sperm populations from individual ejaculates. Using the SDS/citric acid method, RNA was recovered from both fresh and frozen-thawed motile spermatozoa. Sperm RNA were used as templates in reverse transcription-polymerase chain reaction (RT-PCR), in an attempt to identify partial RNA transcripts of a highly conserved region within the alpha-1C (pore-forming) subunit of L-type voltage-dependent calcium channels from 11 individual donors. Control reactions employed primers derived from the human glyceraldehyde-3-phosphate dehydrogenase (GAPDH) sequence. In nine of the 11 specimens, gene-specific PCR products were obtained with both the GAPDH and alpha-1C primer pairs. DNA sequencing analysis confirmed that the respective spliced transcripts were amplified. The two cases in which no amplification was obtained were attributed to reduced RNA yield. These data are consistent with results from in-situ RT-PCR of rat testis sections indicating that the testis-specific calcium channel of that species was expressed uniformly in all stages of the germinal epithelium, including mature spermatozoa.
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PMID:L-type voltage-dependent calcium channel alpha-1C subunit mRNA is present in ejaculated human spermatozoa. 1065 54

Chlorinated antifertility compounds are known to inhibit glycolysis of spermatozoa as they reside in the epididymis but new compounds need to be evaluated that retain antifertility action but do not exhibit side-effects. In this study, two known antifertility agents and a related compound were compared for their inhibition of rat sperm metabolism and motility in vitro. The dose-dependent inhibition in vitro of the glycolytic enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and triosephosphate isomerase (TPI) of distal cauda epididymal rat spermatozoa by (R)-, (S)- and (R,S)-ornidazole (ORN), (R,S)-alpha-chlorohydrin (ACH) and 1-chloro-3-hydroxypropanone (CHOP) was compared. The direct inhibition of GAPDH by ORN suggests that it inhibits without prior conversion outside the cell but inhibition was not stereo-specific. The GAPDH, but not TPI, activity of spermatozoa incubated with ACH and CHOP was highly correlated with kinematic parameters of spermatozoa incubated in pyruvate- and lactate-free medium. ACH only inhibited the activity of intact spermatozoa and the inhibition was not reversed by washing the particulate sperm fraction after sonication. High concentrations of ACH (100 mmol/L) killed intact rat spermatozoa and decreased the extent of GAPDH inhibition. CHOP, unlike ACH, was an effective inhibitor of both intact and sonicated cells. Pre-CHOP, the dimethylketal precursor of CHOP, and its other hydrolysis product MeOH, were both ineffective in vitro. CHOP and related ketals may be more effective inhibitors of sperm glycolysis than ACH and may prove useful for investigating sperm-specific glycolytic inhibition, a prerequisite for the development of antiglycolytic, post-testicular acting contraceptives.
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PMID:In vitro inhibition of rat cauda epididymal sperm glycolytic enzymes by ornidazole, alpha-chlorohydrin and 1-chloro-3-hydroxypropanone. 1101 86

Numerous studies have documented inhibitory effects of alpha-chlorohydrin (ACH) on glyceraldehyde-3-phosphate dehydrogenase (G3PDH) activity in spermatozoa. A sperm-specific G3PDH isoform has been described. The possibility that ACH may inhibit G3PDH in cell types other than sperm was investigated in this work. In addition, the onset of ACH-induced epididymal toxicity was described. Changes to epididymal histology occurred 6 h following a single dose of ACH (50 mg/kg po) and were confined to the proximal initial segment. By 24 h, no epithelial cells lined the basement membrane of that region. Three h after ACH administration (50 mg/kg po), G3PDH activity was significantly decreased in sperm (85%) as well as in kidney (31%), liver (49%), and epididymis (35%). Enzyme activity remained inhibited at 6 and 24 h. G3PDH was immunolocalized in the epididymis and staining was highest in the efferent ducts and initial segment as well as in smooth muscle. Since G3PDH is a microtubule-associated protein and microtubule-dependent endocytosis occurs in the epididymis, beta-tubulin was also immunolocalized. beta-tubulin densely stained the apical region of initial segment and caput epithelial cells. Disruption of beta-tubulin immunostaining correlated with the localization and onset of the lesion. Co-localization of G3PDH and beta-tubulin immunostaining was not observed although both antibodies most densely stained the initial segment. Our data indicate that histologic changes to the proximal initial segment of the epididymis occur rapidly, but subsequent to G3PDH inhibition. Moreover, ACH inhibition of G3PDH is not confined to sperm, although the sperm enzyme is most sensitive to inhibition.
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PMID:alpha-Chlorohydrin inhibits glyceraldehyde-3-phosphate dehydrogenase in multiple organs as well as in sperm. 1139 99

Cytochrome p450 aromatase (p450arom) is a key enzyme responsible for the irreversible transformation of androgens into estrogens. In the present study, we have analysed the ability of human ejaculated spermatozoa to produce estrogens and for that purpose we have looked for the expression of specific aromatase transcript and protein. We have confirmed the presence of p450arom transcript in all normospermic purified samples by nested PCR. The sequence of PCR products from purified spermatozoa shares 98% identity with published human p450arom sequence. Using a semi-quantitative approach, we have observed in immotile sperm a significant decrease (28%) of the aromatase/glyceraldehyde-3-phosphate dehydrogenase ratio compared with the motile sperm fraction. On Western blot with a monoclonal antibody directed against aromatase, we have detected two bands (53 and 49 kDa) in microsome preparations from purified spermatozoa. In total protein extracts of purified spermatozoa (with and without cytoplasmic droplets), we have only found the aromatase as a 49 kDa band with a stronger intensity when cytoplasmic droplets are present. Moreover, the band seems to be weaker in immotile spermatozoa (with and without cytoplasmic droplets). Our data demonstrate the expression of aromatase both in terms of mRNA and protein in each sample of human purified spermatozoa and in addition, our results suggest that aromatase could be concerned with the acquisition of sperm motility.
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PMID:Expression of aromatase in human ejaculated spermatozoa: a putative marker of motility. 1260 87

The mammalian testis serves two main functions: production of spermatozoa and synthesis of steroids; among them, estrogens are the end products obtained from the irreversible transformation of androgens by aromatase. The aromatase is encoded by a single gene (cyp19) in humans which contains 18 exons, 9 of them being translated. In rat the aromatase activity is mainly located in Sertoli cells of immature animals and then in Leydig cells of adults. Moreover rat germ cells represent an additional source of estrogens: the amount of P450arom transcript is 3-fold higher in pachytene spermatocytes (PS) compared to gonocytes or round spermatids (RS); conversely, aromatase activity is more intense in haploid cells. Male germ cells of mice, bank vole, bear and monkey express also aromatase. In man besides Leydig cells, we have shown the presence of a biologically active aromatase and of estrogen receptors in ejaculated spermatozoa and in immature germ cells. Concerning aromatase, a 30% decrease of the amount of mRNA is observed in immotile compared to motile sperm fraction from the same sample; moreover the aromatase activity is also diminished of 34%. In asthenoteratozoospermic and teratozoospermic patients the aromatase gene expression is decreased by 67 and 52%, respectively when compared to normospermic controls. Statistical analyses between the sperm morphology and the aromatase/GAPDH ratio have revealed a high degree of correlation (r=-0.64) between the ratio and the percentage of abnormal spermatozoa (especially microcephaly and acrosmome malformations). Alterations of sperm number and motility have been described in men genetically deficient in aromatase, which together with our data, suggest a likely role for aromatase/estrogens in the acquisition of sperm motility. Therefore besides gonadotrophins and testosterone, estrogens produced locally should be considered as a physiologically relevant hormone involved in the regulation of spermatogenesis and spermiogenesis.
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PMID:Estrogens: a new player in spermatogenesis. 1829 17

The mammalian testis is a complex organ which produces spermatozoa and synthesizes steroids. The transformation of androgens into estrogens is catalyzed by aromatase, an enzymatic complex encoded by a single copy-gene (cyp19) which contains 18 exons, 9 of them being translated. In man besides Leydig cells, we have demonstrated the existence of a biologically active aromatase in immature germ cells and in ejaculated spermatozoa. In addition the presence of estrogen receptors (ERalpha and ERss) in immature germ cells and in spermatozoa has been reported. Concerning aromatase, a 30% decrease of the amount of mRNA is observed in immotile compared to motile sperm fraction from the same sample. In asthenoteratozoospermic, teratozoospermic and asthenozoospermic patients, the aromatase gene expression is decreased respectively by 67%, 52% and 44%, when compared to normospermic controls. Statistical analyses between the sperm morphology and the aromatase/GAPDH ratio have revealed a high degree of correlation (r=-0.64) between that ratio and the percentage of abnormal spermatozoa (especially microcephaly). In men genetically deficient in aromatase diminutions of sperm number and motility have been published. Therefore besides gonadotrophins and testosterone, estrogens are likely playing a relevant role in spermiogenesis and human male gamete maturation.
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PMID:Aromatase and estrogens in man reproduction: a review and latest advances. 1861 33

Boar spermatozoa contain isoforms of both glyceraldehyde 3-phosphate dehydrogenase (GAPDH, EC 1.2.1.12) and pyruvate kinase (PK, EC 2.7.1.40). The sperm-specific forms, GAPDH-S and PK-S, are tightly bound to cell structures. By immunofluorescence microscopy GAPDH-S and PK-S were localised in the principal piece of the boar sperm flagellum as well as in the acrosomal region of the sperm head and at the head-midpiece junction. The midpiece of the flagellum, however, contains isoforms of GAPDH and PK that were only recognised by antibodies against somatic GAPDH and PK, respectively, but not by the antibodies against GAPDH-S and PK-S. In sections of boar testis, GAPDH-S and PK-S were first detected in elongating spermatids when both the developing flagellum and the head were labelled with antibodies against GAPDH-S and PK-S. In contrast, antibodies against rabbit muscle GAPDH and PK labelled all developmental stages of germ cells and also neighbouring contractile cells. Thus, the structure-bound sperm-specific enzymes, GAPDH-S and PK-S, appeared only late in spermatogenesis simultaneously with the development of the structures to which they are bound. Anchoring glycolytic enzymes to structures in these mitochondria-free regions may secure ATP-production for both motility and acrosome function.
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PMID:Expression and compartmentalisation of the glycolytic enzymes GAPDH and pyruvate kinase in boar spermatogenesis. 1867 19

In most mammalian species the aromatase is encoded by a single gene (cyp19), which contains 18 exons, 9 of them being translated. In adult rats, together with Leydig cells germ cells represent an additional source of estrogens. The amount of P450arom transcript is threefold higher in pachytene spermatocytes compared to younger cells (spermatogonia-preleptotene spermatocyte) or round spermatids; conversely, aromatase activity is more intense in haploid cells. In man besides Leydig cells, we have shown the presence of a biologically active aromatase and of estrogen receptors (ERalpha and ERss) in immature germ cells and ejaculated spermatozoa. Concerning aromatase, a 30% decrease of the amount of mRNA is observed in immotile compared to motile sperm fraction from the same sample; moreover, the aromatase activity is diminished. We have amplified aromatase mRNA by RT-real time PCR in spermatozoa from asthenospermic, teratospermic, and asthenoteratospermic men and recorded respectively 44, 52, and 67% decreases of the amount of transcripts as compared to controls. Statistical analyses between the sperm morphology and the aromatase/GAPDH ratio have revealed a high degree of correlation (r = -0.64) with the percentage of abnormal spermatozoa (especially microcephaly and acrosome malformations). Alterations of sperm number and motility have been described in men genetically deficient in aromatase, which together with our data, suggest a likely role for aromatase/estrogens in the acquisition of sperm motility. Therefore besides gonadotrophins and testosterone, estrogens produced locally should be considered as a physiologically relevant hormone involved in the regulation of mammalian spermatogenesis.
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PMID:Mammalian sperm quality and aromatase expression. 1926 95

Sperm glyceraldehyde-3-phosphate dehydrogenase has been shown to be a successful target for a non-hormonal contraceptive approach, but the agents tested to date have had unacceptable side effects. Obtaining the structure of the sperm-specific isoform to allow rational inhibitor design has therefore been a goal for a number of years but has proved intractable because of the insoluble nature of both native and recombinant protein. We have obtained soluble recombinant sperm glyceraldehyde-3-phosphate dehydrogenase as a heterotetramer with the Escherichia coli glyceraldehyde-3-phosphate dehydrogenase in a ratio of 1:3 and have solved the structure of the heterotetramer which we believe represents a novel strategy for structure determination of an insoluble protein. A structure was also obtained where glyceraldehyde 3-phosphate binds in the P(s) pocket in the active site of the sperm enzyme subunit in the presence of NAD. Modeling and comparison of the structures of human somatic and sperm-specific glyceraldehyde-3-phosphate dehydrogenase revealed few differences at the active site and hence rebut the long presumed structural specificity of 3-chlorolactaldehyde for the sperm isoform. The contraceptive activity of alpha-chlorohydrin and its apparent specificity for the sperm isoform in vivo are likely to be due to differences in metabolism to 3-chlorolactaldehyde in spermatozoa and somatic cells. However, further detailed analysis of the sperm glyceraldehyde-3-phosphate dehydrogenase structure revealed sites in the enzyme that do show significant difference compared with published somatic glyceraldehyde-3-phosphate dehydrogenase structures that could be exploited by structure-based drug design to identify leads for novel male contraceptives.
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PMID:Structure of insoluble rat sperm glyceraldehyde-3-phosphate dehydrogenase (GAPDH) via heterotetramer formation with Escherichia coli GAPDH reveals target for contraceptive design. 1954 19

In most mammalian species aromatase is encoded by a single gene (Cyp19), which contains 18 exons, nine of them being translated. In man, the presence of a biologically active aromatase and oestrogen receptors (ERalpha and ERbeta) has been reported in Leydig cells, and also in immature germ cells and ejaculated spermatozoa. Concerning aromatase, the amount of transcript and enzymatic activity are decreased in immotile compared with motile sperm. We have amplified aromatase mRNA by real-time polymerase chain reaction in spermatozoa from asthenospermic, teratospermic and asthenoteratospermic men and recorded, respectively, 44, 52 and 67 per cent decreases of the amount of transcripts compared with fertile donors. A high degree of correlation (r = -0.64) between the abnormal spermatozoa (especially microcephaly and acrosome malformations) and aromatase/GAPDH transcript ratio has been observed. Idiopathic infertility is a wide health problem and no treatment is currently available. In humans, even if the role of oestrogens in spermatogenesis is still a matter of debate, the observations of decreased sperm number and motility in men genetically deficient in aromatase, together with our data and those reported in the literature, may suggest a role for aromatase/oestrogens not only during the development and maintenance of spermatogenesis but also in the final maturation of spermatozoa.
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PMID:Aromatase, oestrogens and human male reproduction. 2040 70


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