Linked Life Data

21671586

Source:http://linkedlifedata.com/resource/pubmed/id/21671586

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
29
pubmed:dateCreated
2011-7-19
pubmed:abstractText
The structure of the flavin-dependent alkanesulfonate monooxygenase (SsuD) exists as a TIM-barrel structure with an insertion region located over the active site that contains a conserved arginine (Arg297) residue present in all SsuD homologues. Substitution of Arg297 with alanine (R297A SsuD) or lysine (R297K SsuD) was performed to determine the functional role of this conserved residue in SsuD catalysis. While the more conservative R297K SsuD possessed a lower k(cat)/K(m) value (0.04 ± 0.01 ?M(-1) min(-1)) relative to wild-type (1.17 ± 0.22 ?M(-1) min(-1)), there was no activity observed with the R297A SsuD variant. Each of the arginine variants had similar K(d) values for flavin binding as wild-type SsuD (0.32 ± 0.15 ?M), but there was no measurable binding of octanesulfonate. The low levels of activity for the R297A and R297K SsuD variants correlated with the absence of any detectable C4a-(peroxy)flavin formation in stopped-flow kinetic studies. Single-turnover experiments were performed in the presence of SsuE to evaluate both the reductive and oxidative half-reaction. With wild-type SsuD a lag phase is observed following the reductive half-reaction by SsuE that represents flavin transfer or conformational changes associated with the binding of substrates. Evaluation of the Arg297 SsuD variants in the presence of SsuE showed no lag phase following reduction by SsuE, and the flavin was oxidized immediately following the reductive half-reaction. These results corresponded with a lack of detectable changes in the proteolytic susceptibility of R297A and R297K SsuD in the presence of reduced flavin and/or octanesulfonate, signifying the absence of a conformational change in these variants with the substitution of Arg297.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jul
pubmed:issn
1520-4995
pubmed:author
pubmed:copyrightInfo
© 2011 American Chemical Society
pubmed:issnType
Electronic
pubmed:day
26
pubmed:volume
50
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
6469-77
pubmed:meshHeading
pubmed:year
2011
pubmed:articleTitle
Functional role of a conserved arginine residue located on a mobile loop of alkanesulfonate monooxygenase.
pubmed:affiliation
Department of Chemistry and Biochemistry, Auburn University, Auburn, Alabama 36849, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, Non-P.H.S.