Linked Life Data

8873618

Source:http://linkedlifedata.com/resource/pubmed/id/8873618

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
41
pubmed:dateCreated
1996-11-27
pubmed:abstractText
O-Acetylserine sulfhydrylase (OASS) is a pyridoxal 5'-phosphate dependent enzyme that catalyzes a beta-replacement reaction forming L-cysteine and acetate from O-acetyl-L-serine (OAS) and sulfide. The pyridoxal 5'-phosphate (PLP) is bound at the active site in Schiff base linkage with a lysine. In the present study, the Schiff base lysine was identified as lysine 42, and its role in the OASS reaction was determined by changing it to alanine using site-directed mutagenesis. K42A-OASS is isolated as an external aldimine with methionine or leucine and shows no reaction with the natural substrates. Apo-K42A-OASS can be reconstituted with PLP, suggesting that K42 is not necessary for cofactor binding and formation of the external Schiff base. The apo-K42A-OASS, reconstituted with PLP, shows slow formation of the external aldimine but does not form the alpha-aminoacrylate intermediate on addition of OAS, suggesting that K42 is involved in the abstraction of the alpha-proton in the beta-elimination reaction. The external aldimines formed upon addition of L-Ala or L-Ser are stable and represent a tautomer that absorbs maximally at 420 nm, while L-Cys gives a tautomeric form of the external aldimine that absorbs at 330 nm, and is also seen in the overall reaction after addition of primary amines to the assay system. The use of a small primary amine such as ethylamine or bromoethylamine in the assay system leads to the initial formation of an internal (gamma-thialysine) or external (ethylamine) aldimine followed by the slow formation of the alpha-aminoacrylate intermediate on addition of OAS. Activity could not be fully recovered, and only a single turnover is observed. Data suggest a significant rate enhancement resulting from the presence of K42 for transimination and general base catalysis.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
15
pubmed:volume
35
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
13485-93
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed-meshheading:8873618-Alanine, pubmed-meshheading:8873618-Amino Acid Sequence, pubmed-meshheading:8873618-Base Sequence, pubmed-meshheading:8873618-Binding Sites, pubmed-meshheading:8873618-Catalysis, pubmed-meshheading:8873618-Cysteine Synthase, pubmed-meshheading:8873618-DNA Primers, pubmed-meshheading:8873618-Escherichia coli, pubmed-meshheading:8873618-Imines, pubmed-meshheading:8873618-Kinetics, pubmed-meshheading:8873618-Lysine, pubmed-meshheading:8873618-Molecular Sequence Data, pubmed-meshheading:8873618-Molecular Structure, pubmed-meshheading:8873618-Mutagenesis, Site-Directed, pubmed-meshheading:8873618-Pyridoxal Phosphate, pubmed-meshheading:8873618-Recombinant Proteins, pubmed-meshheading:8873618-Salmonella typhimurium, pubmed-meshheading:8873618-Schiff Bases, pubmed-meshheading:8873618-Spectrophotometry
pubmed:year
1996
pubmed:articleTitle
A change in the internal aldimine lysine (K42) in O-acetylserine sulfhydrylase to alanine indicates its importance in transimination and as a general base catalyst.
pubmed:affiliation
Department of Biochemistry and Molecular Biology, University of North Texas Health Science Center at Fort Worth 76107-2699, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S., Research Support, Non-U.S. Gov't