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Article: Alanine scan mutagenesis of the switch I domain of the Caulobacter crescentus CgtA protein reveals critical amino acids required for in vivo function

TitleAlanine scan mutagenesis of the switch I domain of the Caulobacter crescentus CgtA protein reveals critical amino acids required for in vivo function
Authors
KeywordsSpecies Index: Bacteria (Microorganisms)
Caulobacter Vibrioides
Negibacteria
Issue Date2001
PublisherBlackwell Publishing Ltd. The Journal's web site is located at http://www.blackwellpublishing.com/journals/MMI
Citation
Molecular Microbiology, 2001, v. 39 n. 4, p. 924-934 How to Cite?
AbstractThe Caulobacter crescentus CgtA protein is a member of the Obg/GTP1 subfamily of monomeric GTP-binding proteins. In vitro, CgtA displays moderate affinity for both GDP and GTP and displays rapid exchange rate constants for either nucleotide, indicating that the guanine nucleotide-binding and exchange properties of CgtA are different from those of the well-characterized Ras-like GTP-binding proteins. The Obg/GTP1 proteins share sequence similarity along the putative effector-binding domain. In this study, we examined the functional consequences of altering amino acid residues within this conserved domain, and identified that T193 was critical for CgtA function. The in vitro binding, exchange and GTP hydrolysis of the T192A, T193A and T192AT193A mutant proteins was examined using fluorescent guanine nucleotide analogues (mant-GDP and mant-GTP). Substitution of either T192 and/or T193 for alanine modestly reduced binding to GDP and significantly reduced the binding affinity for GTP. Furthermore, the T193A mutant protein was more severely impaired for binding GTP than the T192A mutant. The T193A mutation appeared to account solely for the impaired GTP binding of the T192AT193A double mutation. This is the first report that demonstrates that a confirmed defect in guanine nucleotide binding and GTP hydrolysis of an Obg-like protein results in the lack of function in vivo.
Persistent Identifierhttp://hdl.handle.net/10722/91121
ISSN
2015 Impact Factor: 3.761
2015 SCImago Journal Rankings: 2.978
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorLin, Ben_HK
dc.contributor.authorSkidmore, JMen_HK
dc.contributor.authorBhatt, Aen_HK
dc.contributor.authorPfeffer, SMen_HK
dc.contributor.authorPawloski, Len_HK
dc.contributor.authorMaddock, JRen_HK
dc.date.accessioned2010-09-17T10:13:20Z-
dc.date.available2010-09-17T10:13:20Z-
dc.date.issued2001en_HK
dc.identifier.citationMolecular Microbiology, 2001, v. 39 n. 4, p. 924-934en_HK
dc.identifier.issn0950-382Xen_HK
dc.identifier.urihttp://hdl.handle.net/10722/91121-
dc.description.abstractThe Caulobacter crescentus CgtA protein is a member of the Obg/GTP1 subfamily of monomeric GTP-binding proteins. In vitro, CgtA displays moderate affinity for both GDP and GTP and displays rapid exchange rate constants for either nucleotide, indicating that the guanine nucleotide-binding and exchange properties of CgtA are different from those of the well-characterized Ras-like GTP-binding proteins. The Obg/GTP1 proteins share sequence similarity along the putative effector-binding domain. In this study, we examined the functional consequences of altering amino acid residues within this conserved domain, and identified that T193 was critical for CgtA function. The in vitro binding, exchange and GTP hydrolysis of the T192A, T193A and T192AT193A mutant proteins was examined using fluorescent guanine nucleotide analogues (mant-GDP and mant-GTP). Substitution of either T192 and/or T193 for alanine modestly reduced binding to GDP and significantly reduced the binding affinity for GTP. Furthermore, the T193A mutant protein was more severely impaired for binding GTP than the T192A mutant. The T193A mutation appeared to account solely for the impaired GTP binding of the T192AT193A double mutation. This is the first report that demonstrates that a confirmed defect in guanine nucleotide binding and GTP hydrolysis of an Obg-like protein results in the lack of function in vivo.en_HK
dc.languageengen_HK
dc.publisherBlackwell Publishing Ltd. The Journal's web site is located at http://www.blackwellpublishing.com/journals/MMIen_HK
dc.relation.ispartofMolecular Microbiologyen_HK
dc.subjectSpecies Index: Bacteria (Microorganisms)en_HK
dc.subjectCaulobacter Vibrioidesen_HK
dc.subjectNegibacteriaen_HK
dc.titleAlanine scan mutagenesis of the switch I domain of the Caulobacter crescentus CgtA protein reveals critical amino acids required for in vivo functionen_HK
dc.typeArticleen_HK
dc.identifier.emailLin, B:blin@hku.hken_HK
dc.description.naturelink_to_subscribed_fulltext-
dc.identifier.doi10.1046/j.1365-2958.2001.02285.xen_HK
dc.identifier.pmid11251813-
dc.identifier.scopuseid_2-s2.0-0035119263en_HK
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-0035119263&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume39en_HK
dc.identifier.issue4en_HK
dc.identifier.spage924en_HK
dc.identifier.epage934en_HK
dc.identifier.isiWOS:000167237500009-

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