Structure of LacY with an α-substituted galactoside: Connecting the binding site to the protonation site
The X-ray crystal structure of a conformationally constrained mutant of theEscherichia colilactose permease (the LacY double-Trp mutant Gly-46→Trp/Gly-262→Trp) with boundp-nitrophenyl-α-D-galactopyranoside (α-NPG), a high-affinity lactose analog, is described. With the exception of Glu-126 (helix IV...
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| Veröffentlicht in: | Proceedings of the National Academy of Sciences - PNAS 2015-07, Vol.112 (29), p.9004-9009 |
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| container_title | Proceedings of the National Academy of Sciences - PNAS |
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| creator | Kumar, Hemant Finer-Moore, Janet S. Kaback, H. Ronald Stroud, Robert M. |
| description | The X-ray crystal structure of a conformationally constrained mutant of theEscherichia colilactose permease (the LacY double-Trp mutant Gly-46→Trp/Gly-262→Trp) with boundp-nitrophenyl-α-D-galactopyranoside (α-NPG), a high-affinity lactose analog, is described. With the exception of Glu-126 (helix IV), side chains Trp-151 (helix V), Glu-269 (helix VIII), Arg-144 (helix V), His-322 (helix X), and Asn-272 (helix VIII) interact directly with the galactopyranosyl ring of α-NPG to provide specificity, as indicated by biochemical studies and shown directly by X-ray crystallography. In contrast, Phe-20, Met-23, and Phe-27 (helix I) are within van der Waals distance of the benzyl moiety of the analog and thereby increase binding affinity nonspecifically. Thus, the specificity of LacY for sugar is determined solely by side-chain interactions with the galactopyranosyl ring, whereas affinity is increased by nonspecific hydrophobic interactions with the anomeric substitue. |
| doi_str_mv | 10.1073/pnas.1509854112 |
| format | Article |
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Ronald ; Stroud, Robert M.</creator><creatorcontrib>Kumar, Hemant ; Finer-Moore, Janet S. ; Kaback, H. Ronald ; Stroud, Robert M.</creatorcontrib><description>The X-ray crystal structure of a conformationally constrained mutant of theEscherichia colilactose permease (the LacY double-Trp mutant Gly-46→Trp/Gly-262→Trp) with boundp-nitrophenyl-α-D-galactopyranoside (α-NPG), a high-affinity lactose analog, is described. With the exception of Glu-126 (helix IV), side chains Trp-151 (helix V), Glu-269 (helix VIII), Arg-144 (helix V), His-322 (helix X), and Asn-272 (helix VIII) interact directly with the galactopyranosyl ring of α-NPG to provide specificity, as indicated by biochemical studies and shown directly by X-ray crystallography. In contrast, Phe-20, Met-23, and Phe-27 (helix I) are within van der Waals distance of the benzyl moiety of the analog and thereby increase binding affinity nonspecifically. Thus, the specificity of LacY for sugar is determined solely by side-chain interactions with the galactopyranosyl ring, whereas affinity is increased by nonspecific hydrophobic interactions with the anomeric substitue.</description><identifier>ISSN: 0027-8424</identifier><identifier>EISSN: 1091-6490</identifier><identifier>DOI: 10.1073/pnas.1509854112</identifier><identifier>PMID: 26157133</identifier><language>eng</language><publisher>United States: National Academy of Sciences</publisher><subject>Binding Sites ; Biological Sciences ; Crystallography, X-Ray ; Escherichia coli - enzymology ; Escherichia coli Proteins - chemistry ; Escherichia coli Proteins - metabolism ; hydrogen bonding ; hydrophobic bonding ; hydrophobicity ; lactose ; Ligands ; Models, Molecular ; Monosaccharide Transport Proteins - chemistry ; Monosaccharide Transport Proteins - metabolism ; mutants ; Nitrophenylgalactosides - metabolism ; Protein Structure, Secondary ; Protons ; Static Electricity ; Substrate Specificity ; Symporters - chemistry ; Symporters - metabolism</subject><ispartof>Proceedings of the National Academy of Sciences - PNAS, 2015-07, Vol.112 (29), p.9004-9009</ispartof><rights>Volumes 1–89 and 106–112, copyright as a collective work only; author(s) retains copyright to individual articles</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c542t-d462b7677605db2963e44bb12d23727ffee6658ac45e33122cd939aa9e014e513</citedby><cites>FETCH-LOGICAL-c542t-d462b7677605db2963e44bb12d23727ffee6658ac45e33122cd939aa9e014e513</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Uhttp://www.pnas.org/content/112/29.cover.gif</thumbnail><linktopdf>$$Uhttps://www.jstor.org/stable/pdf/26466100$$EPDF$$P50$$Gjstor$$H</linktopdf><linktohtml>$$Uhttps://www.jstor.org/stable/26466100$$EHTML$$P50$$Gjstor$$H</linktohtml><link.rule.ids>230,314,723,776,780,799,881,27901,27902,53766,53768,57992,58225</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/26157133$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kumar, Hemant</creatorcontrib><creatorcontrib>Finer-Moore, Janet S.</creatorcontrib><creatorcontrib>Kaback, H. Ronald</creatorcontrib><creatorcontrib>Stroud, Robert M.</creatorcontrib><title>Structure of LacY with an α-substituted galactoside: Connecting the binding site to the protonation site</title><title>Proceedings of the National Academy of Sciences - PNAS</title><addtitle>Proc Natl Acad Sci U S A</addtitle><description>The X-ray crystal structure of a conformationally constrained mutant of theEscherichia colilactose permease (the LacY double-Trp mutant Gly-46→Trp/Gly-262→Trp) with boundp-nitrophenyl-α-D-galactopyranoside (α-NPG), a high-affinity lactose analog, is described. With the exception of Glu-126 (helix IV), side chains Trp-151 (helix V), Glu-269 (helix VIII), Arg-144 (helix V), His-322 (helix X), and Asn-272 (helix VIII) interact directly with the galactopyranosyl ring of α-NPG to provide specificity, as indicated by biochemical studies and shown directly by X-ray crystallography. In contrast, Phe-20, Met-23, and Phe-27 (helix I) are within van der Waals distance of the benzyl moiety of the analog and thereby increase binding affinity nonspecifically. Thus, the specificity of LacY for sugar is determined solely by side-chain interactions with the galactopyranosyl ring, whereas affinity is increased by nonspecific hydrophobic interactions with the anomeric substitue.</description><subject>Binding Sites</subject><subject>Biological Sciences</subject><subject>Crystallography, X-Ray</subject><subject>Escherichia coli - enzymology</subject><subject>Escherichia coli Proteins - chemistry</subject><subject>Escherichia coli Proteins - metabolism</subject><subject>hydrogen bonding</subject><subject>hydrophobic bonding</subject><subject>hydrophobicity</subject><subject>lactose</subject><subject>Ligands</subject><subject>Models, Molecular</subject><subject>Monosaccharide Transport Proteins - chemistry</subject><subject>Monosaccharide Transport Proteins - metabolism</subject><subject>mutants</subject><subject>Nitrophenylgalactosides - metabolism</subject><subject>Protein Structure, Secondary</subject><subject>Protons</subject><subject>Static Electricity</subject><subject>Substrate Specificity</subject><subject>Symporters - chemistry</subject><subject>Symporters - metabolism</subject><issn>0027-8424</issn><issn>1091-6490</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqNkT1v1EAQhlcRiByBOhXEZRonM7Nf3iZSFBFAOokCKKhW6_U6ceSzL7vrRPws_gi_CZ_uOJKOaop53kczehk7RjhD0Px8Pbh0hhJMJQUiHbAFgsFSCQMv2AKAdFkJEofsdUp3AGBkBa_YISmUGjlfMPE1x8nnKYZibIul8z-Kxy7fFm4ofv8q01Sn3OUph6a4cb3zeUxdE96wl63rU3i7m0fs-_WHb1efyuWXj5-vLpell4Jy2QhFtVZaK5BNTUbxIERdIzXENem2DUEpWTkvZOAciXxjuHHOBEARJPIjdrH1rqd6FRofhhxdb9exW7n4046us883Q3drb8YHKyRqIpgFpztBHO-nkLJddcmHvndDGKdksUJdcTTqP1BlKm4IuZzR8y3q45hSDO3-IgS7qcVuarH_apkT758-suf_9jADJztgk9zrkCwZawDETLzbEncpj_GJQSiFAPwPcqKcdw</recordid><startdate>20150721</startdate><enddate>20150721</enddate><creator>Kumar, Hemant</creator><creator>Finer-Moore, Janet S.</creator><creator>Kaback, H. 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Ronald ; Stroud, Robert M.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c542t-d462b7677605db2963e44bb12d23727ffee6658ac45e33122cd939aa9e014e513</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Binding Sites</topic><topic>Biological Sciences</topic><topic>Crystallography, X-Ray</topic><topic>Escherichia coli - enzymology</topic><topic>Escherichia coli Proteins - chemistry</topic><topic>Escherichia coli Proteins - metabolism</topic><topic>hydrogen bonding</topic><topic>hydrophobic bonding</topic><topic>hydrophobicity</topic><topic>lactose</topic><topic>Ligands</topic><topic>Models, Molecular</topic><topic>Monosaccharide Transport Proteins - chemistry</topic><topic>Monosaccharide Transport Proteins - metabolism</topic><topic>mutants</topic><topic>Nitrophenylgalactosides - metabolism</topic><topic>Protein Structure, Secondary</topic><topic>Protons</topic><topic>Static Electricity</topic><topic>Substrate Specificity</topic><topic>Symporters - chemistry</topic><topic>Symporters - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kumar, Hemant</creatorcontrib><creatorcontrib>Finer-Moore, Janet S.</creatorcontrib><creatorcontrib>Kaback, H. 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Ronald</au><au>Stroud, Robert M.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Structure of LacY with an α-substituted galactoside: Connecting the binding site to the protonation site</atitle><jtitle>Proceedings of the National Academy of Sciences - PNAS</jtitle><addtitle>Proc Natl Acad Sci U S A</addtitle><date>2015-07-21</date><risdate>2015</risdate><volume>112</volume><issue>29</issue><spage>9004</spage><epage>9009</epage><pages>9004-9009</pages><issn>0027-8424</issn><eissn>1091-6490</eissn><abstract>The X-ray crystal structure of a conformationally constrained mutant of theEscherichia colilactose permease (the LacY double-Trp mutant Gly-46→Trp/Gly-262→Trp) with boundp-nitrophenyl-α-D-galactopyranoside (α-NPG), a high-affinity lactose analog, is described. With the exception of Glu-126 (helix IV), side chains Trp-151 (helix V), Glu-269 (helix VIII), Arg-144 (helix V), His-322 (helix X), and Asn-272 (helix VIII) interact directly with the galactopyranosyl ring of α-NPG to provide specificity, as indicated by biochemical studies and shown directly by X-ray crystallography. In contrast, Phe-20, Met-23, and Phe-27 (helix I) are within van der Waals distance of the benzyl moiety of the analog and thereby increase binding affinity nonspecifically. Thus, the specificity of LacY for sugar is determined solely by side-chain interactions with the galactopyranosyl ring, whereas affinity is increased by nonspecific hydrophobic interactions with the anomeric substitue.</abstract><cop>United States</cop><pub>National Academy of Sciences</pub><pmid>26157133</pmid><doi>10.1073/pnas.1509854112</doi><tpages>6</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Binding Sites Biological Sciences Crystallography, X-Ray Escherichia coli - enzymology Escherichia coli Proteins - chemistry Escherichia coli Proteins - metabolism hydrogen bonding hydrophobic bonding hydrophobicity lactose Ligands Models, Molecular Monosaccharide Transport Proteins - chemistry Monosaccharide Transport Proteins - metabolism mutants Nitrophenylgalactosides - metabolism Protein Structure, Secondary Protons Static Electricity Substrate Specificity Symporters - chemistry Symporters - metabolism |
| title | Structure of LacY with an α-substituted galactoside: Connecting the binding site to the protonation site |
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