The N-terminal cysteine of human asparagine synthetase is essential for glutamine-dependent activity

• Published in: The Journal of biological chemistry Vol. 264; no. 33; pp. 19475 - 19477
• Main Authors: VAN HEEKE, G, SCHUSTER, S. M
• Format: Journal Article
• Language: English
• Published: Bethesda, MD American Society for Biochemistry and Molecular Biology 25.11.1989
• Subjects: Alanine; Amidophosphoribosyltransferase - genetics; Amino Acid Sequence; Analytical, structural and metabolic biochemistry; Aspartate-Ammonia Ligase - genetics; Aspartate-Ammonia Ligase - metabolism; Bacillus subtilis - enzymology; Bacillus subtilis - genetics; Base Sequence; Biological and medical sciences; Cloning, Molecular; Cysteine; Enzymes and enzyme inhibitors; Escherichia coli - enzymology; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Glutamine - metabolism; Humans; Ligases; Ligases - genetics; man; Molecular Sequence Data; Mutation; Oligonucleotide Probes; Plasmids; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Sequence Homology, Nucleic Acid; Enzyme; N terminal aminoacid
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/s0021-9258(19)47138-x

Site-specific mutagenesis was used to replace the N-terminal cysteine in human asparagine synthetase by an alanine. The mutant enzyme was expressed in the yeast Saccharomyces cerevisiae, and the asparagine synthetase activity was analyzed in vitro. The mutation resulted in the loss of the glutamine-dependent asparagine synthetase activity, while the ammonia-dependent activity remained unaffected. These results confirm the existence of a glutamine amidotransfer domain with an N-terminal cysteine essential for the glutamine-dependent asparagine synthetase activity.