Linked Life Data

21616082

Source:http://linkedlifedata.com/resource/pubmed/id/21616082

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
2011-6-24
pubmed:databankReference
pubmed:abstractText
The oxidative catabolism of uric acid produces 5-hydroxyisourate (HIU), which is further degraded to (S)-allantoin by two enzymes, HIU hydrolase and 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline decarboxylase. The intermediates of the latter two reactions, HIU and 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline, are unstable in solution and decay nonstereospecifically to allantoin. In addition, nonenzymatic racemization of allantoin has been shown to occur at physiological pH. Since the further breakdown of allantoin is catalyzed by allantoinase, an enzyme that is specific for (S)-allantoin, an allantoin racemase is necessary for complete and efficient catabolism of uric acid. In this work, we characterize the structure and activity of allantoin racemase from Klebsiella pneumoniae (KpHpxA). In addition to an unliganded structure solved using selenomethionyl single-wavelength anomalous dispersion, structures of C79S/C184S KpHpxA in complex with allantoin and with 5-acetylhydantoin are presented. These structures reveal several important features of the active site including an oxyanion hole and a polar binding pocket that interacts with the ureido tail of allantoin and serves to control the orientation of the hydantoin ring. The ability of KpHpxA to interconvert the (R)- and (S)-enantiomers of allantoin is demonstrated, and analysis of the steady-state kinetics of KpHpxA yielded a k(cat)/K(m) of 6.0 × 10(5) M(-1) s(-1). Mutation of either of the active-site cysteines, Cys79 or Cys184, to serine inactivates this enzyme. The data presented provide new insights into the activity and substrate specificity of this enzyme and enable us to propose a mechanism for catalysis that is consistent with the two-base mechanism observed in other members of the aspartate/glutamate family.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jul
pubmed:issn
1089-8638
pubmed:author
pubmed:copyrightInfo
Copyright © 2011 Elsevier Ltd. All rights reserved.
pubmed:issnType
Electronic
pubmed:day
15
pubmed:volume
410
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
447-60
pubmed:meshHeading
pubmed-meshheading:21616082-Allantoin, pubmed-meshheading:21616082-Amino Acid Substitution, pubmed-meshheading:21616082-Bacterial Proteins, pubmed-meshheading:21616082-Binding Sites, pubmed-meshheading:21616082-Biocatalysis, pubmed-meshheading:21616082-Catalytic Domain, pubmed-meshheading:21616082-Crystallography, X-Ray, pubmed-meshheading:21616082-Cysteine, pubmed-meshheading:21616082-Humans, pubmed-meshheading:21616082-Kinetics, pubmed-meshheading:21616082-Klebsiella Infections, pubmed-meshheading:21616082-Klebsiella pneumoniae, pubmed-meshheading:21616082-Models, Molecular, pubmed-meshheading:21616082-Mutation, pubmed-meshheading:21616082-Protein Multimerization, pubmed-meshheading:21616082-Protein Structure, Quaternary, pubmed-meshheading:21616082-Protein Structure, Secondary, pubmed-meshheading:21616082-Racemases and Epimerases, pubmed-meshheading:21616082-Serine, pubmed-meshheading:21616082-Stereoisomerism, pubmed-meshheading:21616082-Substrate Specificity
pubmed:year
2011
pubmed:articleTitle
Characterization of the structure and function of Klebsiella pneumoniae allantoin racemase.
pubmed:affiliation
Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY 14853, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, Non-U.S. Gov't, Research Support, N.I.H., Extramural