Linked Life Data

18498176

Source:http://linkedlifedata.com/resource/pubmed/id/18498176

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
24
pubmed:dateCreated
2008-6-10
pubmed:databankReference
pubmed:abstractText
A crystal structure of serine acetyltransferase (SAT) with cysteine bound in the serine subsite of the active site shows that both H154 and H189 are within hydrogen-bonding distance to the cysteine thiol [Olsen, L. R., Huang, B., Vetting, M. W., and Roderick, S. L. (2004) Biochemistry 43, 6013 -6019]. In addition, H154 is in an apparent dyad linkage with D139. The structure suggests that H154 is the most likely catalytic general base and that H189 and D139 may also play important roles during the catalytic reaction. Site-directed mutagenesis was performed to mutate each of these three residues to Asn, one at a time. The V1/Et value of all of the single mutant enzymes decreased, with the largest decrease (approximately 1240-fold) exhibited by the H154N mutant enzyme. Mutation of both histidines, H154N/H189N, gave a V1/Et approximately 23700-fold lower than that of the wild-type enzyme. An increase in K Ser was observed for the H189N, D139N, and H154N/H189N mutant enzymes, while the H154N mutant enzyme gave an 8-fold decrease in K Ser. For all three single mutant enzymes, V1/Et and V1/K Ser Et decrease at low pH and give a pKa of about 7, while the V1/Et of the double mutant enzyme was pH independent. The solvent deuterium kinetic isotope effects on V 1 and V1/K Ser decreased compared to wild type for the H154N mutant enzyme and increased for the H189N mutant enzyme but was about the same as that of wild type for D139N and H154N/H189N. Data suggest that H154, H189, and D139 play different catalytic roles for SAT. H154 likely serves as a general base, accepting a proton from the beta-hydroxyl of serine as the tetrahedral intermediate is formed upon nucleophilic attack on the thioester carbonyl of acetyl-CoA. However, activity is not completely lost upon elimination of H154, and thus, H189 may be able to serve as a backup general base at a lower efficiency compared to H154; it also aids in binding and orienting the serine substrate. Aspartate 139, in dyad linkage with H154, likely facilitates catalysis by increasing the basicity of H154.
pubmed:grant
pubmed:commentsCorrections
http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-1427014, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-15147185, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-15313214, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-15333931, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-15581365, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-15581568, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-21524, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-359557, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-502857, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-5332668, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-5550822, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-6855597, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-7481807, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-8332530, http://linkedlifedata.com/resource/pubmed/commentcorrection/18498176-9890979
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
1520-4995
pubmed:author
pubmed:issnType
Electronic
pubmed:day
17
pubmed:volume
47
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
6322-8
pubmed:dateRevised
2010-9-21
pubmed:meshHeading
pubmed:year
2008
pubmed:articleTitle
Roles of histidines 154 and 189 and aspartate 139 in the active site of serine acetyltransferase from Haemophilus influenzae.
pubmed:affiliation
Department of Chemistry and Biochemistry, University of Oklahoma, 620 Parrington Oval, Norman, Oklahoma 73019, USA.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't