Determination of amino acid sequences involved in the processing of the ARG5/ARG6 precursor in Saccharomyces cerevisiae.
In Saccharomyces cerevisiae, the ARG5/ARG6 locus encodes both acetylglutamate kinase and acetylglutamyl-phosphate reductase, localized in the mitochondria. Genetic analysis, determination of the nucleotide sequence of the ARG5/ARG6 gene and identification of the transcript indicate that it encodes a single translation product containing two enzyme activities. However, analysis of cellular extracts revealed that the activities are completely separable. In this work, we define different domains in the ARG5/ARG6 polypeptide; a mitochondrial target sequence and the acetylglutamate-kinase and acetylglutamyl-phosphate-reductase domains. We show that deletions in the N-terminal end of the protein and point mutations in the junction region between the acetylglutamate-kinase and acetylglutamyl-phosphate-reductase domains lead to the accumulation of large precursor. Our data support the idea that import of the ARG5/ARG6 precursor into the mitochondria is required for its processing into two mature enzymes.
Boonchird
C
C
Erfelijkheidsleer en Microbiologie, Vrije Universiteit, Brussels, Belgium.
Messenguy
F
F
Dubois
E
E
D000445:Aldehyde Oxidoreductases / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D004792:Enzyme Precursors / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005798:Genes, Bacterial; D008928:Mitochondria / Q000201:enzymology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015345:Oligonucleotide Probes; D010770:Phosphotransferases / Q000235:genetics*; D017851:Phosphotransferases (Carboxyl Group Acceptor); D010957:Plasmids; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid
D004792:Enzyme Precursors; D015345:Oligonucleotide Probes; D021382:Protein Sorting Signals; D000445:Aldehyde Oxidoreductases; C018722:N-acetyl-gamma-glutamyl-phosphate reductase; D010770:Phosphotransferases; D017851:Phosphotransferases (Carboxyl Group Acceptor); C018721:acetylglutamate kinase