Reengineering Rate-Limiting, Millisecond Enzyme Motions by Introduction of an Unnatural Amino Acid
Identifieur interne : 000057 ( Pmc/Curation ); précédent : 000056; suivant : 000058Reengineering Rate-Limiting, Millisecond Enzyme Motions by Introduction of an Unnatural Amino Acid
Auteurs : Eric D. Watt [États-Unis] ; Ivan Rivalta [États-Unis] ; Sean K. Whittier [États-Unis] ; Victor S. Batista [États-Unis] ; J. Patrick Loria [États-Unis]Source :
- Biophysical Journal [ 0006-3495 ] ; 2011.
Abstract
Rate-limiting millisecond motions in wild-type (WT) Ribonuclease A (RNase A) are modulated by histidine 48. Here, we incorporate an unnatural amino acid, thia-methylimidazole, at this site (H48C-4MI) to investigate the effects of a single residue on protein motions over multiple timescales and on enzyme catalytic turnover. Molecular dynamics simulations reveal that H48C-4MI retains some crucial WT-like hydrogen bonding interactions but the extent of protein-wide correlated motions in the nanosecond regime is decreased relative to WT. NMR Carr-Purcell-Meiboom-Gill relaxation dispersion experiments demonstrate that millisecond conformational motions in H48C-4MI are present over a similar pH range compared to WT. Furthermore, incorporation of this nonnatural amino acid allows retention of WT-like catalytic activity over the full pH range. These studies demonstrate that the complexity of the protein energy landscape during the catalytic cycle can be maintained using unnatural amino acids, which may prove useful in enzyme design efforts.
Url:
DOI: 10.1016/j.bpj.2011.05.039
PubMed: 21767494
PubMed Central: 3136797
Links toward previous steps (curation, corpus...)
- to stream Pmc, to step Corpus: Pour aller vers cette notice dans l'étape Curation :000057
Links to Exploration step
PMC:3136797Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Reengineering Rate-Limiting, Millisecond Enzyme Motions by Introduction of an Unnatural Amino Acid</title><author><name sortKey="Watt, Eric D" sort="Watt, Eric D" uniqKey="Watt E" first="Eric D." last="Watt">Eric D. Watt</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Rivalta, Ivan" sort="Rivalta, Ivan" uniqKey="Rivalta I" first="Ivan" last="Rivalta">Ivan Rivalta</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Whittier, Sean K" sort="Whittier, Sean K" uniqKey="Whittier S" first="Sean K." last="Whittier">Sean K. Whittier</name><affiliation wicri:level="2"><nlm:aff id="aff2">Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Molecular Biophysics and Biochemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Batista, Victor S" sort="Batista, Victor S" uniqKey="Batista V" first="Victor S." last="Batista">Victor S. Batista</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Loria, J Patrick" sort="Loria, J Patrick" uniqKey="Loria J" first="J. Patrick" last="Loria">J. Patrick Loria</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation><affiliation wicri:level="2"><nlm:aff id="aff2">Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Molecular Biophysics and Biochemistry, Yale University, New Haven</wicri:cityArea></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">21767494</idno><idno type="pmc">3136797</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3136797</idno><idno type="RBID">PMC:3136797</idno><idno type="doi">10.1016/j.bpj.2011.05.039</idno><date when="2011">2011</date><idno type="wicri:Area/Pmc/Corpus">000057</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000057</idno><idno type="wicri:Area/Pmc/Curation">000057</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Curation">000057</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Reengineering Rate-Limiting, Millisecond Enzyme Motions by Introduction of an Unnatural Amino Acid</title><author><name sortKey="Watt, Eric D" sort="Watt, Eric D" uniqKey="Watt E" first="Eric D." last="Watt">Eric D. Watt</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Rivalta, Ivan" sort="Rivalta, Ivan" uniqKey="Rivalta I" first="Ivan" last="Rivalta">Ivan Rivalta</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Whittier, Sean K" sort="Whittier, Sean K" uniqKey="Whittier S" first="Sean K." last="Whittier">Sean K. Whittier</name><affiliation wicri:level="2"><nlm:aff id="aff2">Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Molecular Biophysics and Biochemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Batista, Victor S" sort="Batista, Victor S" uniqKey="Batista V" first="Victor S." last="Batista">Victor S. Batista</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation></author><author><name sortKey="Loria, J Patrick" sort="Loria, J Patrick" uniqKey="Loria J" first="J. Patrick" last="Loria">J. Patrick Loria</name><affiliation wicri:level="2"><nlm:aff id="aff1">Department of Chemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Chemistry, Yale University, New Haven</wicri:cityArea></affiliation><affiliation wicri:level="2"><nlm:aff id="aff2">Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Connecticut</region></placeName><wicri:cityArea>Department of Molecular Biophysics and Biochemistry, Yale University, New Haven</wicri:cityArea></affiliation></author></analytic><series><title level="j">Biophysical Journal</title><idno type="ISSN">0006-3495</idno><idno type="eISSN">1542-0086</idno><imprint><date when="2011">2011</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>Rate-limiting millisecond motions in wild-type (WT) Ribonuclease A (RNase A) are modulated by histidine 48. Here, we incorporate an unnatural amino acid, thia-methylimidazole, at this site (H48C-4MI) to investigate the effects of a single residue on protein motions over multiple timescales and on enzyme catalytic turnover. Molecular dynamics simulations reveal that H48C-4MI retains some crucial WT-like hydrogen bonding interactions but the extent of protein-wide correlated motions in the nanosecond regime is decreased relative to WT. NMR Carr-Purcell-Meiboom-Gill relaxation dispersion experiments demonstrate that millisecond conformational motions in H48C-4MI are present over a similar pH range compared to WT. Furthermore, incorporation of this nonnatural amino acid allows retention of WT-like catalytic activity over the full pH range. These studies demonstrate that the complexity of the protein energy landscape during the catalytic cycle can be maintained using unnatural amino acids, which may prove useful in enzyme design efforts.</p></div></front></TEI><pmc article-type="research-article"><pmc-comment>The publisher of this article does not allow downloading of the full text in XML form.</pmc-comment>
<front><journal-meta><journal-id journal-id-type="nlm-ta">Biophys J</journal-id><journal-title-group><journal-title>Biophysical Journal</journal-title></journal-title-group><issn pub-type="ppub">0006-3495</issn><issn pub-type="epub">1542-0086</issn><publisher><publisher-name>The Biophysical Society</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">21767494</article-id><article-id pub-id-type="pmc">3136797</article-id><article-id pub-id-type="publisher-id">BPJ2923</article-id><article-id pub-id-type="doi">10.1016/j.bpj.2011.05.039</article-id><article-categories><subj-group subj-group-type="heading"><subject>Protein</subject></subj-group></article-categories><title-group><article-title>Reengineering Rate-Limiting, Millisecond Enzyme Motions by Introduction of an Unnatural Amino Acid</article-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>Watt</surname><given-names>Eric D.</given-names></name><xref rid="aff1" ref-type="aff">†</xref></contrib><contrib contrib-type="author"><name><surname>Rivalta</surname><given-names>Ivan</given-names></name><xref rid="aff1" ref-type="aff">†</xref></contrib><contrib contrib-type="author"><name><surname>Whittier</surname><given-names>Sean K.</given-names></name><xref rid="aff2" ref-type="aff">‡</xref></contrib><contrib contrib-type="author"><name><surname>Batista</surname><given-names>Victor S.</given-names></name><xref rid="aff1" ref-type="aff">†</xref></contrib><contrib contrib-type="author"><name><surname>Loria</surname><given-names>J. Patrick</given-names></name><email>patrick.loria@yale.edu</email><xref rid="aff1" ref-type="aff">†</xref><xref rid="aff2" ref-type="aff">‡</xref><xref rid="cor1" ref-type="corresp">∗</xref></contrib></contrib-group><aff id="aff1"><label>†</label>Department of Chemistry, Yale University, New Haven, Connecticut</aff><aff id="aff2"><label>‡</label>Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut</aff><author-notes><corresp id="cor1"><label>∗</label>Corresponding author <email>patrick.loria@yale.edu</email></corresp></author-notes><pub-date pub-type="pmc-release"><day>20</day><month>7</month><year>2011</year></pub-date><pmc-comment> PMC Release delay is 0 months and 0 days and was based on the
<pub-date pub-type="ppub"><day>20</day><month>7</month><year>2011</year></pub-date><volume>101</volume><issue>2</issue><fpage>411</fpage><lpage>420</lpage><history><date date-type="received"><day>28</day><month>3</month><year>2011</year></date><date date-type="accepted"><day>19</day><month>5</month><year>2011</year></date></history><permissions><copyright-statement>© 2011 by the Biophysical Society.</copyright-statement><copyright-year>2011</copyright-year><copyright-holder>Biophysical Society</copyright-holder></permissions><abstract><p>Rate-limiting millisecond motions in wild-type (WT) Ribonuclease A (RNase A) are modulated by histidine 48. Here, we incorporate an unnatural amino acid, thia-methylimidazole, at this site (H48C-4MI) to investigate the effects of a single residue on protein motions over multiple timescales and on enzyme catalytic turnover. Molecular dynamics simulations reveal that H48C-4MI retains some crucial WT-like hydrogen bonding interactions but the extent of protein-wide correlated motions in the nanosecond regime is decreased relative to WT. NMR Carr-Purcell-Meiboom-Gill relaxation dispersion experiments demonstrate that millisecond conformational motions in H48C-4MI are present over a similar pH range compared to WT. Furthermore, incorporation of this nonnatural amino acid allows retention of WT-like catalytic activity over the full pH range. These studies demonstrate that the complexity of the protein energy landscape during the catalytic cycle can be maintained using unnatural amino acids, which may prove useful in enzyme design efforts.</p></abstract></article-meta></front></pmc></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Wicri/Lorraine/explor/InforLorV4/Data/Pmc/Curation
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 000057 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Pmc/Curation/biblio.hfd -nk 000057 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Wicri/Lorraine
|area= InforLorV4
|flux= Pmc
|étape= Curation
|type= RBID
|clé= PMC:3136797
|texte= Reengineering Rate-Limiting, Millisecond Enzyme Motions by Introduction of an Unnatural Amino Acid
}}
Pour générer des pages wiki
HfdIndexSelect -h $EXPLOR_AREA/Data/Pmc/Curation/RBID.i -Sk "pubmed:21767494" \
| HfdSelect -Kh $EXPLOR_AREA/Data/Pmc/Curation/biblio.hfd \
| NlmPubMed2Wicri -a InforLorV4
| This area was generated with Dilib version V0.6.33. |  |