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Title:A Characterization of Aspartyl Peptidases in Salmonella Typhimurium
Author(s):Lassy, Rachel Anne Larsen
Doctoral Committee Chair(s):Miller, Charles G.
Department / Program:Microbiology
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Chemistry, Biochemistry
Abstract:Few of the peptidases previously characterized from Salmonella typhimurium are capable of hydrolyzing aspartyl peptides. Peptidase E, an aspartyl specific dipeptidase, was the first member of a new family of peptide hydrolases that now includes peptidases from other proteobacteria and from two eukaryotes. It is shown that Peptidase E from Xenopus laevis has the same substrate specificity as Peptidase E from S. typhimurium, indicating that in this family of enzymes both the catalytic and the substrate specificity have been conserved. An alignment of the amino acid sequences of the Peptidase E family allowed for potentially important residues to be identified. Through site-directed mutagenesis of the S. typhimurium pepE, it was proposed that this enzyme is a serine hydrolase that utilizes a catalytic triad of serine, aspartate, and histidine. In extracts of a pepE strain, two additional Asp-X hydrolases were detected. The identity of each of these enzymes as well as evidence of a third Asp-X hydrolase are presented in this thesis. Two of these enzymes are shown most rapidly hydrolyze isoaspartyl peptides (also called beta-aspartyl peptides). Of the two isoaspartyl peptidases, one is a homolog of IadA from E. coli and the other is described for the first time in this work and is encoded by a previously unknown open reading frame called ybiK in E. coli. The ybiK gene product is a threonine hydrolase that is a member of the Ntn (N-terminal nucleophile) family of enzymes. It is synthesized as a 32 kD polypeptide that dimerizes and undergoes processing, presumably autocatalytic, into an active form, which is a 60 kD heterotetramer. A strain of S. typhimurium lacking all of the broad specificity peptidases, as well as the aspartyl peptidases described here is still capable of growth on Asp-Leu as a leucine source. This growth is attributed to the remaining peptidase. This peptidase is specific for alpha-aspartyl peptides, has a native molecular mass of 70 kD, and has the unusual property of requiring manganese for activity.
Issue Date:2000
Description:111 p.
Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2000.
Other Identifier(s):(MiAaPQ)AAI9955640
Date Available in IDEALS:2015-09-28
Date Deposited:2000

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