Prolyl oligopeptidase catalysis Reactions with thiono substrates reveal substrate-induced conformational change to be the rate-limiting step
Prolyl oligopeptidase, a member of the new family of serine proteases, exhibits significant mechanistic differences compared with the enzymes of the chymotrypsin and subtilisin families. Our kinetic study using the thiono substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide suggests that the...
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| Veröffentlicht in: | FEBS letters 1993-05, Vol.322 (3), p.227-230 |
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| creator | Polgár, László Kollát, Emma Hollósi, Miklós |
| description | Prolyl oligopeptidase, a member of the new family of serine proteases, exhibits significant mechanistic differences compared with the enzymes of the chymotrypsin and subtilisin families. Our kinetic study using the thiono substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide suggests that the putative oxyanion binding site is important in prolyl oligopeptidase catalysis, although to a lesser extent than in the chymotrypsin-and subtilisin-catalyzed reactions. By using another thiono substrate, benzyloxycarbonyl-Gly[CS-NH]Pro-2-naphthylamide, it is demonstrated that the distant S2P2 hydrogen bond (formed between the S2 subsite and P2 peptide residue) makes a greater contribution to catalysis than does stabilization by the oxyanion binding site involved directly in the bond cleavage. In contrast to the reactions catalyzed by chymotrypsin and subtilisin, no kinetic deuterium isotope effect is apparent in the acylation of prolyl oligopeptidase measured either with the specific benzyloxycarbonyl-Gly-Pro-2-naphthylamide, or with the very poor substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide. This indicates that the rate-limiting conformational change is induced by the substrate. |
| doi_str_mv | 10.1016/0014-5793(93)81575-K |
| format | Article |
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Our kinetic study using the thiono substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide suggests that the putative oxyanion binding site is important in prolyl oligopeptidase catalysis, although to a lesser extent than in the chymotrypsin-and subtilisin-catalyzed reactions. By using another thiono substrate, benzyloxycarbonyl-Gly[CS-NH]Pro-2-naphthylamide, it is demonstrated that the distant S2P2 hydrogen bond (formed between the S2 subsite and P2 peptide residue) makes a greater contribution to catalysis than does stabilization by the oxyanion binding site involved directly in the bond cleavage. In contrast to the reactions catalyzed by chymotrypsin and subtilisin, no kinetic deuterium isotope effect is apparent in the acylation of prolyl oligopeptidase measured either with the specific benzyloxycarbonyl-Gly-Pro-2-naphthylamide, or with the very poor substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide. This indicates that the rate-limiting conformational change is induced by the substrate.</description><identifier>ISSN: 0014-5793</identifier><identifier>EISSN: 1873-3468</identifier><identifier>DOI: 10.1016/0014-5793(93)81575-K</identifier><identifier>PMID: 8486154</identifier><language>eng</language><publisher>England: Elsevier B.V</publisher><subject>2-naphthylamide ; Animals ; benzyloxycarbonyl ; Binding Sites ; Boc ; Conformational change ; dithioerythritol ; DTE ; Enzyme mechanism ; Glyt and Prot ; Hydrogen Bonding ; Hydrogen-Ion Concentration ; Kinetics ; Nap ; Oxygen - metabolism ; Protein Conformation ; Rate-limiting step ; represent Gly and Pro, respectively, containing a sulfur atom in place of the carbonyl oxygen ; Serine Endopeptidases - chemistry ; Serine Endopeptidases - metabolism ; Subsite specificity ; Substrate Specificity ; Swine ; t-butyloxycarbonyl ; Thiones - metabolism ; Thiono substrate</subject><ispartof>FEBS letters, 1993-05, Vol.322 (3), p.227-230</ispartof><rights>1993</rights><rights>FEBS Letters 322 (1993) 1873-3468 © 2015 Federation of European Biochemical Societies</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c420K-95c2508170d53e4f7d1d2fe67da1004e79e105a197fc33877857bb4d0adc2acc3</citedby><cites>FETCH-LOGICAL-c420K-95c2508170d53e4f7d1d2fe67da1004e79e105a197fc33877857bb4d0adc2acc3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/0014-5793(93)81575-K$$EHTML$$P50$$Gelsevier$$Hfree_for_read</linktohtml><link.rule.ids>315,781,785,3551,27928,27929,45999</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/8486154$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Polgár, László</creatorcontrib><creatorcontrib>Kollát, Emma</creatorcontrib><creatorcontrib>Hollósi, Miklós</creatorcontrib><title>Prolyl oligopeptidase catalysis Reactions with thiono substrates reveal substrate-induced conformational change to be the rate-limiting step</title><title>FEBS letters</title><addtitle>FEBS Lett</addtitle><description>Prolyl oligopeptidase, a member of the new family of serine proteases, exhibits significant mechanistic differences compared with the enzymes of the chymotrypsin and subtilisin families. Our kinetic study using the thiono substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide suggests that the putative oxyanion binding site is important in prolyl oligopeptidase catalysis, although to a lesser extent than in the chymotrypsin-and subtilisin-catalyzed reactions. By using another thiono substrate, benzyloxycarbonyl-Gly[CS-NH]Pro-2-naphthylamide, it is demonstrated that the distant S2P2 hydrogen bond (formed between the S2 subsite and P2 peptide residue) makes a greater contribution to catalysis than does stabilization by the oxyanion binding site involved directly in the bond cleavage. In contrast to the reactions catalyzed by chymotrypsin and subtilisin, no kinetic deuterium isotope effect is apparent in the acylation of prolyl oligopeptidase measured either with the specific benzyloxycarbonyl-Gly-Pro-2-naphthylamide, or with the very poor substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide. This indicates that the rate-limiting conformational change is induced by the substrate.</description><subject>2-naphthylamide</subject><subject>Animals</subject><subject>benzyloxycarbonyl</subject><subject>Binding Sites</subject><subject>Boc</subject><subject>Conformational change</subject><subject>dithioerythritol</subject><subject>DTE</subject><subject>Enzyme mechanism</subject><subject>Glyt and Prot</subject><subject>Hydrogen Bonding</subject><subject>Hydrogen-Ion Concentration</subject><subject>Kinetics</subject><subject>Nap</subject><subject>Oxygen - metabolism</subject><subject>Protein Conformation</subject><subject>Rate-limiting step</subject><subject>represent Gly and Pro, respectively, containing a sulfur atom in place of the carbonyl oxygen</subject><subject>Serine Endopeptidases - chemistry</subject><subject>Serine Endopeptidases - metabolism</subject><subject>Subsite specificity</subject><subject>Substrate Specificity</subject><subject>Swine</subject><subject>t-butyloxycarbonyl</subject><subject>Thiones - metabolism</subject><subject>Thiono substrate</subject><issn>0014-5793</issn><issn>1873-3468</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1993</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqNUdGO1CAUJUazjqt_oAlPRh-qUGBoX0x0s6NmNtEYfSYUbmcwtFSgu5l_8KOlM5P1zZgQuJx7zoGci9BzSt5QQtdvCaG8ErJlr1r2uqFCimr7AK1oI1nF-Lp5iFb3lMfoSUo_Sbk3tL1AFw1v1lTwFfr9NQZ_8Dh4twsTTNlZnQAbnbU_JJfwN9AmuzAmfOfyHud9qQNOc5dy1BkSjnAL2v9FKjfa2YDFJox9iINe1IVg9nrcAc4Bd2XfAz6SvRtcduMOpwzTU_So1z7Bs_N5iX5srr9ffapuvnz8fPX-pjK8JtuqFaYWpKGSWMGA99JSW_ewllZTQjjIFigRmrayN4w1UjZCdh23RFtTa2PYJXp58p1i-DVDympwyYD3eoQwJyWFrFtR14XIT0QTQ0oRejVFN-h4UJSoZQhqSVgtCauyjkNQ2yJ7cfafuwHsveiceulvTv075-HwX55qc_2hXhoL3rIjujz07mQEJa1bB1El42As6bsIJisb3L9_-gcc3a3f</recordid><startdate>19930517</startdate><enddate>19930517</enddate><creator>Polgár, László</creator><creator>Kollát, Emma</creator><creator>Hollósi, Miklós</creator><general>Elsevier B.V</general><scope>6I.</scope><scope>AAFTH</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>19930517</creationdate><title>Prolyl oligopeptidase catalysis Reactions with thiono substrates reveal substrate-induced conformational change to be the rate-limiting step</title><author>Polgár, László ; Kollát, Emma ; Hollósi, Miklós</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c420K-95c2508170d53e4f7d1d2fe67da1004e79e105a197fc33877857bb4d0adc2acc3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1993</creationdate><topic>2-naphthylamide</topic><topic>Animals</topic><topic>benzyloxycarbonyl</topic><topic>Binding Sites</topic><topic>Boc</topic><topic>Conformational change</topic><topic>dithioerythritol</topic><topic>DTE</topic><topic>Enzyme mechanism</topic><topic>Glyt and Prot</topic><topic>Hydrogen Bonding</topic><topic>Hydrogen-Ion Concentration</topic><topic>Kinetics</topic><topic>Nap</topic><topic>Oxygen - metabolism</topic><topic>Protein Conformation</topic><topic>Rate-limiting step</topic><topic>represent Gly and Pro, respectively, containing a sulfur atom in place of the carbonyl oxygen</topic><topic>Serine Endopeptidases - chemistry</topic><topic>Serine Endopeptidases - metabolism</topic><topic>Subsite specificity</topic><topic>Substrate Specificity</topic><topic>Swine</topic><topic>t-butyloxycarbonyl</topic><topic>Thiones - metabolism</topic><topic>Thiono substrate</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Polgár, László</creatorcontrib><creatorcontrib>Kollát, Emma</creatorcontrib><creatorcontrib>Hollósi, Miklós</creatorcontrib><collection>ScienceDirect Open Access Titles</collection><collection>Elsevier:ScienceDirect:Open Access</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>FEBS letters</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Polgár, László</au><au>Kollát, Emma</au><au>Hollósi, Miklós</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Prolyl oligopeptidase catalysis Reactions with thiono substrates reveal substrate-induced conformational change to be the rate-limiting step</atitle><jtitle>FEBS letters</jtitle><addtitle>FEBS Lett</addtitle><date>1993-05-17</date><risdate>1993</risdate><volume>322</volume><issue>3</issue><spage>227</spage><epage>230</epage><pages>227-230</pages><issn>0014-5793</issn><eissn>1873-3468</eissn><abstract>Prolyl oligopeptidase, a member of the new family of serine proteases, exhibits significant mechanistic differences compared with the enzymes of the chymotrypsin and subtilisin families. Our kinetic study using the thiono substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide suggests that the putative oxyanion binding site is important in prolyl oligopeptidase catalysis, although to a lesser extent than in the chymotrypsin-and subtilisin-catalyzed reactions. By using another thiono substrate, benzyloxycarbonyl-Gly[CS-NH]Pro-2-naphthylamide, it is demonstrated that the distant S2P2 hydrogen bond (formed between the S2 subsite and P2 peptide residue) makes a greater contribution to catalysis than does stabilization by the oxyanion binding site involved directly in the bond cleavage. In contrast to the reactions catalyzed by chymotrypsin and subtilisin, no kinetic deuterium isotope effect is apparent in the acylation of prolyl oligopeptidase measured either with the specific benzyloxycarbonyl-Gly-Pro-2-naphthylamide, or with the very poor substrate, benzyloxycarbonyl-Gly-Pro[CS-NH]-2-naphthylamide. This indicates that the rate-limiting conformational change is induced by the substrate.</abstract><cop>England</cop><pub>Elsevier B.V</pub><pmid>8486154</pmid><doi>10.1016/0014-5793(93)81575-K</doi><tpages>4</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | FEBS letters, 1993-05, Vol.322 (3), p.227-230 |
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| source | MEDLINE; Access via ScienceDirect (Elsevier); EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection |
| subjects | 2-naphthylamide Animals benzyloxycarbonyl Binding Sites Boc Conformational change dithioerythritol DTE Enzyme mechanism Glyt and Prot Hydrogen Bonding Hydrogen-Ion Concentration Kinetics Nap Oxygen - metabolism Protein Conformation Rate-limiting step represent Gly and Pro, respectively, containing a sulfur atom in place of the carbonyl oxygen Serine Endopeptidases - chemistry Serine Endopeptidases - metabolism Subsite specificity Substrate Specificity Swine t-butyloxycarbonyl Thiones - metabolism Thiono substrate |
| title | Prolyl oligopeptidase catalysis Reactions with thiono substrates reveal substrate-induced conformational change to be the rate-limiting step |
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