A polyprotein precursor of two mitochondrial enzymes in Neurospora crassa. Gene structure and precursor processing.
N-Acetylglutamate kinase (AGK) and N-acetyl-gamma-glutamyl-phosphate reductase (AGPR) function as two separate mitochondrial enzymes, but are encoded by a single nuclear gene in several fungi. The Neurospora crassa arg-6 gene encoding these enzymes has been cloned and sequenced, and the enzymes responsible for processing the polyprotein precursor have been identified. The 871-amino acid precursor contains a normal N-terminal mitochondrial targeting sequence, an internal connecting region (approximately 200 amino acids) upstream of the distal reductase domain, and coding regions with N-terminal amino acid sequences identical with those of purified N-acetylglutamate kinase and N-acetyl-gamma-glutamyl-phosphate reductase. Sequence comparisons of the coding regions indicate high levels of conservation between prokaryotic and fungal proteins. Regions suggesting ancestral relationships to N-acetylglutamate synthase and aspartate beta-semialdehyde dehydrogenase have been identified. Both the N-terminal targeting sequence and the connecting region contain consensus sites for cleavage by the mitochondrial processing peptidase and processing enhancing protein. In vitro processing assays with intact mitochondria, solubilized mitochondria, and purified enzymes have shown that the mitochondrial processing peptidase and processing enhancing protein cleave not only the N-terminal mitochondrial targeting sequence but also process the polyprotein precursor into the two mature enzymes.
Gessert
S F
SF
Department of Chemistry and Biochemistry, University of California, Los Angeles 90024-1569.
Kim
J H
JH
Nargang
F E
FE
Weiss
R L
RL
D000445:Aldehyde Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics; D000595:Amino Acid Sequence; D001483:Base Sequence; D005075:Biological Evolution; D002874:Chromosome Mapping; D015825:Chromosomes, Fungal; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003360:Cosmids; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005800:Genes, Fungal; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D009492:Neurospora crassa / Q000201:enzymology* / Q000235:genetics*; D017851:Phosphotransferases (Carboxyl Group Acceptor) / Q000096:biosynthesis* / Q000235:genetics; D010957:Plasmids; D012150:Polymorphism, Restriction Fragment Length; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D014170:Transformation, Genetic
D004792:Enzyme Precursors; D000445:Aldehyde Oxidoreductases; C018722:N-acetyl-gamma-glutamyl-phosphate reductase; D017851:Phosphotransferases (Carboxyl Group Acceptor); C018721:acetylglutamate kinase