Beta-D-galactosidase of rat spermatozoa: subcellular distribution, substrate specificity, and molecular changes during epididymal maturation
In previous studies, we reported that rat epididymal fluid acid beta-D-galactosidase, which optimally cleaves a synthetic substrate (PNP beta-D-galactoside) at pH 3.5, shows maximum activity at pH 6.8 when a glycoprotein is used as a substrate [Skudlarek MD, Tulsiani DRP, Orgebin-Crist M-C. Biochem...
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| Veröffentlicht in: | Biology of reproduction 1993-08, Vol.49 (2), p.204-213 |
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| creator | SKUDLAREK, M. D TULSIANI, D. R. P NAGDAS, S. K ORGEBIN-CRIST, M.-C |
| description | In previous studies, we reported that rat epididymal fluid acid beta-D-galactosidase, which optimally cleaves a synthetic
substrate (PNP beta-D-galactoside) at pH 3.5, shows maximum activity at pH 6.8 when a glycoprotein is used as a substrate
[Skudlarek MD, Tulsiani DRP, Orgebin-Crist M-C. Biochem J 1992; 286: 907-914]. We now describe a similar pH-dependent substrate
preference for rat sperm beta-D-galactosidase. We found that only 10-14% of total beta-D-galactosidase (and other glycosidase)
activity was associated with spermatozoa. The remaining enzyme activities were present in soluble form in the luminal fluid.
When the glycosidase levels were expressed per 10(6) sperm, all enzymes showed a progressive increase in spermatozoa from
the caput to the corpus or proximal cauda followed by a sharp decline in spermatozoa from the distal cauda epididymidis. The
observed decrease in beta-D-galactosidase activity could not be explained by the loss of cytoplasmic droplets (which have
a low enzyme activity relative to spermatozoa) or the presence of inhibitors/activators of the enzyme activity in spermatozoa
from the proximal or distal epididymis. However, we found that the changes in beta-D-galactosidase activity during sperm maturation
in the epididymis were accompanied by changes in the molecular form(s) of the enzyme. Western blot analysis using an antibody
to beta-D-galactosidase showed a progressive processing of the 82-kDa immunoreactive band in caput spermatozoa to an 80-kDa
diffuse band in cauda spermatozoa. The sperm-associated beta-D-galactosidase form(s) does not appear to be due to adsorption
and/or binding of the luminal fluid beta-D-galactosidase, which contained a 97-kDa form in fluid from the caput and two forms,
of 97 kDa and 84 kDa, in corpus and cauda fluids. The observed difference in the molecular forms of the sperm and luminal
fluid was found to be due to differential glycosylation, since de-N-glycosylation of various forms of beta-D-galactosidase
generated a single immunoreactive form of 70 kDa. Subcellular localization studies and assay for the beta-D-galactosidase
activity in the enriched plasma membrane and acrosomal membrane fractions suggested the likelihood that the activity of beta-D-galactosidase
and other glycosidases is present in the acrosome and is readily released during sperm disruption. The evidence suggests that
sperm beta-D-galactosidase may be functional within the acidic environment of the acrosome during sperm maturatio |
| doi_str_mv | 10.1095/biolreprod49.2.204 |
| format | Article |
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substrate (PNP beta-D-galactoside) at pH 3.5, shows maximum activity at pH 6.8 when a glycoprotein is used as a substrate
[Skudlarek MD, Tulsiani DRP, Orgebin-Crist M-C. Biochem J 1992; 286: 907-914]. We now describe a similar pH-dependent substrate
preference for rat sperm beta-D-galactosidase. We found that only 10-14% of total beta-D-galactosidase (and other glycosidase)
activity was associated with spermatozoa. The remaining enzyme activities were present in soluble form in the luminal fluid.
When the glycosidase levels were expressed per 10(6) sperm, all enzymes showed a progressive increase in spermatozoa from
the caput to the corpus or proximal cauda followed by a sharp decline in spermatozoa from the distal cauda epididymidis. The
observed decrease in beta-D-galactosidase activity could not be explained by the loss of cytoplasmic droplets (which have
a low enzyme activity relative to spermatozoa) or the presence of inhibitors/activators of the enzyme activity in spermatozoa
from the proximal or distal epididymis. However, we found that the changes in beta-D-galactosidase activity during sperm maturation
in the epididymis were accompanied by changes in the molecular form(s) of the enzyme. Western blot analysis using an antibody
to beta-D-galactosidase showed a progressive processing of the 82-kDa immunoreactive band in caput spermatozoa to an 80-kDa
diffuse band in cauda spermatozoa. The sperm-associated beta-D-galactosidase form(s) does not appear to be due to adsorption
and/or binding of the luminal fluid beta-D-galactosidase, which contained a 97-kDa form in fluid from the caput and two forms,
of 97 kDa and 84 kDa, in corpus and cauda fluids. The observed difference in the molecular forms of the sperm and luminal
fluid was found to be due to differential glycosylation, since de-N-glycosylation of various forms of beta-D-galactosidase
generated a single immunoreactive form of 70 kDa. Subcellular localization studies and assay for the beta-D-galactosidase
activity in the enriched plasma membrane and acrosomal membrane fractions suggested the likelihood that the activity of beta-D-galactosidase
and other glycosidases is present in the acrosome and is readily released during sperm disruption. The evidence suggests that
sperm beta-D-galactosidase may be functional within the acidic environment of the acrosome during sperm maturation as well
as in the neutral environment of the oviduct after the zona-induced acrosome reaction.</description><identifier>ISSN: 0006-3363</identifier><identifier>EISSN: 1529-7268</identifier><identifier>DOI: 10.1095/biolreprod49.2.204</identifier><identifier>PMID: 8373943</identifier><identifier>CODEN: BIREBV</identifier><language>eng</language><publisher>Madison, WI: Society for the Study of Reproduction</publisher><subject>Animals ; beta-Galactosidase - metabolism ; Biological and medical sciences ; Cell Fractionation ; Centrifugation, Density Gradient ; Cytoplasm - enzymology ; Epididymis - enzymology ; Epididymis - growth & development ; Epididymis - ultrastructure ; Fundamental and applied biological sciences. Psychology ; Glycosylation ; Male ; Mammalian male genital system ; Morphology. Physiology ; Rats ; Rats, Sprague-Dawley ; Spermatozoa - enzymology ; Spermatozoa - ultrastructure ; Subcellular Fractions - enzymology ; Substrate Specificity ; Vertebrates: reproduction</subject><ispartof>Biology of reproduction, 1993-08, Vol.49 (2), p.204-213</ispartof><rights>1993 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=4856295$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/8373943$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>SKUDLAREK, M. D</creatorcontrib><creatorcontrib>TULSIANI, D. R. P</creatorcontrib><creatorcontrib>NAGDAS, S. K</creatorcontrib><creatorcontrib>ORGEBIN-CRIST, M.-C</creatorcontrib><title>Beta-D-galactosidase of rat spermatozoa: subcellular distribution, substrate specificity, and molecular changes during epididymal maturation</title><title>Biology of reproduction</title><addtitle>Biol Reprod</addtitle><description>In previous studies, we reported that rat epididymal fluid acid beta-D-galactosidase, which optimally cleaves a synthetic
substrate (PNP beta-D-galactoside) at pH 3.5, shows maximum activity at pH 6.8 when a glycoprotein is used as a substrate
[Skudlarek MD, Tulsiani DRP, Orgebin-Crist M-C. Biochem J 1992; 286: 907-914]. We now describe a similar pH-dependent substrate
preference for rat sperm beta-D-galactosidase. We found that only 10-14% of total beta-D-galactosidase (and other glycosidase)
activity was associated with spermatozoa. The remaining enzyme activities were present in soluble form in the luminal fluid.
When the glycosidase levels were expressed per 10(6) sperm, all enzymes showed a progressive increase in spermatozoa from
the caput to the corpus or proximal cauda followed by a sharp decline in spermatozoa from the distal cauda epididymidis. The
observed decrease in beta-D-galactosidase activity could not be explained by the loss of cytoplasmic droplets (which have
a low enzyme activity relative to spermatozoa) or the presence of inhibitors/activators of the enzyme activity in spermatozoa
from the proximal or distal epididymis. However, we found that the changes in beta-D-galactosidase activity during sperm maturation
in the epididymis were accompanied by changes in the molecular form(s) of the enzyme. Western blot analysis using an antibody
to beta-D-galactosidase showed a progressive processing of the 82-kDa immunoreactive band in caput spermatozoa to an 80-kDa
diffuse band in cauda spermatozoa. The sperm-associated beta-D-galactosidase form(s) does not appear to be due to adsorption
and/or binding of the luminal fluid beta-D-galactosidase, which contained a 97-kDa form in fluid from the caput and two forms,
of 97 kDa and 84 kDa, in corpus and cauda fluids. The observed difference in the molecular forms of the sperm and luminal
fluid was found to be due to differential glycosylation, since de-N-glycosylation of various forms of beta-D-galactosidase
generated a single immunoreactive form of 70 kDa. Subcellular localization studies and assay for the beta-D-galactosidase
activity in the enriched plasma membrane and acrosomal membrane fractions suggested the likelihood that the activity of beta-D-galactosidase
and other glycosidases is present in the acrosome and is readily released during sperm disruption. The evidence suggests that
sperm beta-D-galactosidase may be functional within the acidic environment of the acrosome during sperm maturation as well
as in the neutral environment of the oviduct after the zona-induced acrosome reaction.</description><subject>Animals</subject><subject>beta-Galactosidase - metabolism</subject><subject>Biological and medical sciences</subject><subject>Cell Fractionation</subject><subject>Centrifugation, Density Gradient</subject><subject>Cytoplasm - enzymology</subject><subject>Epididymis - enzymology</subject><subject>Epididymis - growth & development</subject><subject>Epididymis - ultrastructure</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Glycosylation</subject><subject>Male</subject><subject>Mammalian male genital system</subject><subject>Morphology. Physiology</subject><subject>Rats</subject><subject>Rats, Sprague-Dawley</subject><subject>Spermatozoa - enzymology</subject><subject>Spermatozoa - ultrastructure</subject><subject>Subcellular Fractions - enzymology</subject><subject>Substrate Specificity</subject><subject>Vertebrates: reproduction</subject><issn>0006-3363</issn><issn>1529-7268</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1993</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNo9kd1uEzEQhS0EKqHwAkhIvgCuusHrn82aOyg_rVSJG7heje3ZxMi7DrZXUfoMfWgcGnHjkXW-c-yZIeR1y9Yt0-qD8TEk3KfopF7zNWfyCVm1iutmw7v-KVkxxrpGiE48Jy9y_s1YKwUXF-SiFxuhpViRh89YoPnSbCGALTF7BxlpHGmCQvMe0wQl3kf4SPNiLIawBEjU-VySN0vxcb46KfUKBU8G60dvfTleUZgdnWJA-89idzBvMVO3JD9vKe698-44QaD1haW6a9RL8myEkPHVuV6SX9--_ry-ae5-fL-9_nTX7HinSmN6x3ouuVZ8VE5blEq2SnW2tRo7bkbZ23oaxhlWAHVvjdo4adqeGzBGXJL3j7l1dH8WzGWYfD41BzPGJQ8bpaVksq3gmzO4mAndsE9-gnQczuOr-tuzDtlCGBPM1uf_mOxVV39ZsXeP2M5vdwefcMi18VBDxXA4HKQe-FCXJ_4C_KqSHA</recordid><startdate>19930801</startdate><enddate>19930801</enddate><creator>SKUDLAREK, M. D</creator><creator>TULSIANI, D. R. P</creator><creator>NAGDAS, S. K</creator><creator>ORGEBIN-CRIST, M.-C</creator><general>Society for the Study of Reproduction</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>7X8</scope></search><sort><creationdate>19930801</creationdate><title>Beta-D-galactosidase of rat spermatozoa: subcellular distribution, substrate specificity, and molecular changes during epididymal maturation</title><author>SKUDLAREK, M. D ; TULSIANI, D. R. P ; NAGDAS, S. K ; ORGEBIN-CRIST, M.-C</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-h265t-b8d08242952f5d9ce4541556c1c9e62bf48c2bfb020ef5de98cb57d4b182babb3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1993</creationdate><topic>Animals</topic><topic>beta-Galactosidase - metabolism</topic><topic>Biological and medical sciences</topic><topic>Cell Fractionation</topic><topic>Centrifugation, Density Gradient</topic><topic>Cytoplasm - enzymology</topic><topic>Epididymis - enzymology</topic><topic>Epididymis - growth & development</topic><topic>Epididymis - ultrastructure</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Glycosylation</topic><topic>Male</topic><topic>Mammalian male genital system</topic><topic>Morphology. Physiology</topic><topic>Rats</topic><topic>Rats, Sprague-Dawley</topic><topic>Spermatozoa - enzymology</topic><topic>Spermatozoa - ultrastructure</topic><topic>Subcellular Fractions - enzymology</topic><topic>Substrate Specificity</topic><topic>Vertebrates: reproduction</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>SKUDLAREK, M. D</creatorcontrib><creatorcontrib>TULSIANI, D. R. P</creatorcontrib><creatorcontrib>NAGDAS, S. K</creatorcontrib><creatorcontrib>ORGEBIN-CRIST, M.-C</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>MEDLINE - Academic</collection><jtitle>Biology of reproduction</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>SKUDLAREK, M. D</au><au>TULSIANI, D. R. P</au><au>NAGDAS, S. K</au><au>ORGEBIN-CRIST, M.-C</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Beta-D-galactosidase of rat spermatozoa: subcellular distribution, substrate specificity, and molecular changes during epididymal maturation</atitle><jtitle>Biology of reproduction</jtitle><addtitle>Biol Reprod</addtitle><date>1993-08-01</date><risdate>1993</risdate><volume>49</volume><issue>2</issue><spage>204</spage><epage>213</epage><pages>204-213</pages><issn>0006-3363</issn><eissn>1529-7268</eissn><coden>BIREBV</coden><abstract>In previous studies, we reported that rat epididymal fluid acid beta-D-galactosidase, which optimally cleaves a synthetic
substrate (PNP beta-D-galactoside) at pH 3.5, shows maximum activity at pH 6.8 when a glycoprotein is used as a substrate
[Skudlarek MD, Tulsiani DRP, Orgebin-Crist M-C. Biochem J 1992; 286: 907-914]. We now describe a similar pH-dependent substrate
preference for rat sperm beta-D-galactosidase. We found that only 10-14% of total beta-D-galactosidase (and other glycosidase)
activity was associated with spermatozoa. The remaining enzyme activities were present in soluble form in the luminal fluid.
When the glycosidase levels were expressed per 10(6) sperm, all enzymes showed a progressive increase in spermatozoa from
the caput to the corpus or proximal cauda followed by a sharp decline in spermatozoa from the distal cauda epididymidis. The
observed decrease in beta-D-galactosidase activity could not be explained by the loss of cytoplasmic droplets (which have
a low enzyme activity relative to spermatozoa) or the presence of inhibitors/activators of the enzyme activity in spermatozoa
from the proximal or distal epididymis. However, we found that the changes in beta-D-galactosidase activity during sperm maturation
in the epididymis were accompanied by changes in the molecular form(s) of the enzyme. Western blot analysis using an antibody
to beta-D-galactosidase showed a progressive processing of the 82-kDa immunoreactive band in caput spermatozoa to an 80-kDa
diffuse band in cauda spermatozoa. The sperm-associated beta-D-galactosidase form(s) does not appear to be due to adsorption
and/or binding of the luminal fluid beta-D-galactosidase, which contained a 97-kDa form in fluid from the caput and two forms,
of 97 kDa and 84 kDa, in corpus and cauda fluids. The observed difference in the molecular forms of the sperm and luminal
fluid was found to be due to differential glycosylation, since de-N-glycosylation of various forms of beta-D-galactosidase
generated a single immunoreactive form of 70 kDa. Subcellular localization studies and assay for the beta-D-galactosidase
activity in the enriched plasma membrane and acrosomal membrane fractions suggested the likelihood that the activity of beta-D-galactosidase
and other glycosidases is present in the acrosome and is readily released during sperm disruption. The evidence suggests that
sperm beta-D-galactosidase may be functional within the acidic environment of the acrosome during sperm maturation as well
as in the neutral environment of the oviduct after the zona-induced acrosome reaction.</abstract><cop>Madison, WI</cop><pub>Society for the Study of Reproduction</pub><pmid>8373943</pmid><doi>10.1095/biolreprod49.2.204</doi><tpages>10</tpages></addata></record> |
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| ispartof | Biology of reproduction, 1993-08, Vol.49 (2), p.204-213 |
| issn | 0006-3363 1529-7268 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_75944041 |
| source | MEDLINE; EZB Electronic Journals Library |
| subjects | Animals beta-Galactosidase - metabolism Biological and medical sciences Cell Fractionation Centrifugation, Density Gradient Cytoplasm - enzymology Epididymis - enzymology Epididymis - growth & development Epididymis - ultrastructure Fundamental and applied biological sciences. Psychology Glycosylation Male Mammalian male genital system Morphology. Physiology Rats Rats, Sprague-Dawley Spermatozoa - enzymology Spermatozoa - ultrastructure Subcellular Fractions - enzymology Substrate Specificity Vertebrates: reproduction |
| title | Beta-D-galactosidase of rat spermatozoa: subcellular distribution, substrate specificity, and molecular changes during epididymal maturation |
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