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Type: Journal article
Title: Molecular recognition in a post-translational modification of exceptional specificity
Author: Chapman-Smith, A.
Morris, T.
Wallace, J.
Cronan Jr, J.
Citation: Journal of Biological Chemistry, 1999; 274(3):1449-1457
Issue Date: 1999
ISSN: 0021-9258
Statement of
Anne Chapman-Smith, Timothy W. Morris, John C. Wallace, and John E. Cronan, Jr.
Abstract: We have used localized mutagenesis of the biotin domain of the Escherichia coli biotin carboxyl carrier protein coupled with a genetic selection to identify regions of the domain having a role in interactions with the modifying enzyme, biotin protein ligase. We purified several singly substituted mutant biotin domains that showed reduced biotinylation in vivo and characterized these proteins in vitro. This approach has allowed us to distinguish putative biotin protein ligase interaction mutations from structurally defective proteins. Two mutant proteins with glutamate to lysine substitutions (at residues 119 or 147) behaved as authentic ligase interaction mutants. The E119K protein was virtually inactive as a substrate for biotin protein ligase, whereas the E147K protein could be biotinylated, albeit poorly. Neither substitution affected the overall structure of the domain, assayed by disulfide dimer formation and trypsin resistance. Substitutions of the highly conserved glycine residues at positions 133 and 143 or at a key hydrophobic core residue, Val-146, gave structurally unstable proteins.
Keywords: Escherichia coli
Acetyl-CoA Carboxylase
Carbon-Nitrogen Ligases
Carrier Proteins
Recombinant Fusion Proteins
Amino Acid Substitution
Mutagenesis, Site-Directed
Protein Processing, Post-Translational
Binding Sites
Amino Acid Sequence
Protein Binding
Protein Folding
Structure-Activity Relationship
Mutation, Missense
Models, Chemical
Models, Molecular
Molecular Sequence Data
Fatty Acid Synthase, Type II
Rights: © 1999 by The American Society for Biochemistry and Molecular Biology, Inc.
DOI: 10.1074/jbc.274.3.1449
Appears in Collections:Aurora harvest 7
Biochemistry publications

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