Structure of the trifunctional trp-1 gene from Neurospora crassa and its aberrant expression in Escherichia coli

• Published in: Journal of molecular and applied genetics Vol. 2; no. 1; p. 83
• Main Authors: Schechtman, M G, Yanofsky, C
• Format: Journal Article
• Language: English
• Published: United States 1983
• Subjects: Base Sequence; Biological Evolution; Carboxy-Lyases - genetics; Cloning, Molecular; Codon; Escherichia coli - genetics; Gene Expression Regulation; Genes; Indole-3-Glycerol-Phosphate Synthase - genetics; Neurospora - genetics; Neurospora crassa - genetics; Protein Biosynthesis; RNA, Messenger - genetics; Tryptophan - genetics
• ISSN: 0271-6801

The trifunctional trp-1 gene from Neurospora crassa was cloned by complementation of a phosphoribosylanthranilate isomerase-deficient mutant of E. coli. A 2.7-kb DNA sequence containing trp-1 was determined. Homology of the deduced trp-1 polypeptide sequence to the corresponding E. coli proteins is striking; the order of functional domains within trp-1 is NH2-glutamine amidotransferase-indoleglycerolphosphate synthase-phosphoribosylanthranilate isomerase-COOH (NH2-trpG-trpC-trpF-COOH). Whereas trpF complementing activity can be detected in E. coli, trpC activity is absent. It is likely that translation of trp-1 does not proceed from the proper start site in E. coli; the carboxy terminal portion of the trp-1 polypeptide may be the only portion synthesized. Fusion of a bacterial amino terminus and ribosome binding site to the trp-1 coding region results in expression of trpC as well as trpF activity in E. coli. The locations of several startpoints for trp-1 mRNA synthesis were determined by the S1 nuclease mapping technique. DNA immediately 5' to the trp-1 transcription initiation region does not possess a sequence resembling the canonical TATAAA of eukaryotes.