Please use this identifier to cite or link to this item: http://hdl.handle.net/2440/11269
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Type: Journal article
Title: Polymorphism of the yeast pyruvate carboxylase 2 gene and protein: Effects on biotinylation
Author: Val, D.
Chapman-Smith, A.
Walker, M.
Cronan, J.
Wallace, J.
Citation: Biochemical Journal, 1995; 312(3):817-825
Publisher: The Biochemical Society, London
Issue Date: 1995
ISSN: 0264-6021
1470-8728
Abstract: In Saccharomyces cerevisiae there are two isoenzymes of pyruvate carboxylase (Pyc) encoded by separate genes designated PYC1 and PYC2. We report the isolation and sequencing of a PYC2 gene, and the localization of both genes on the physical map of S. cerevisiae. Comparison with the previously reported sequence [Stucka, Dequin, Salmon and Gancedo (1991) Mol. Gen. Genet. 229, 307-315] revealed significant differences within the open reading frame. The most notable difference was near the 3' end, where we found a single base deletion reducing the open reading frame by 15 bases. We have confirmed the C-terminus of Pyc2 encoded by the gene isolated here by expressing and purifying an 86-amino-acid biotin-domain peptide. In addition, we investigated the effects of the two changes in the Pyc2 biotin domain (K1155R substitution and Q1178P/five-amino-acid extension) on the extent of biotinylation in vivo by Escherichia coli biotin ligase, and compared the biotinylation of peptides containing these changes with that of two different-length Pyc1 biotin-domain peptides. The K1155R substitution had very little effect on biotinylation, but the five-amino-acid C-terminal extension to Pyc2 and the N-terminal extension to Pycl both improved biotinylation in vivo.
Keywords: Escherichia coli; Saccharomyces cerevisiae; Biotin; Isoenzymes; Pyruvate Carboxylase; DNA, Fungal; Chromosome Mapping; Sequence Analysis; Mutagenesis; Binding Sites; Amino Acid Sequence; Base Sequence; Sequence Homology; Polymorphism, Genetic; Genes, Fungal; Molecular Sequence Data
RMID: 0030004420
DOI: 10.1042/bj3120817
Appears in Collections:Biochemistry publications

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