ISSN:
1617-4623
Keywords:
Schizosaccharomyces pombe
;
Histidine
;
Shuttle vector
;
Cloning
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract A DNA fragment which carries the his3 gene of Schizosaccharomyes pombe has been isolated and characterized for use as a selectable marker in transformations. The his3 gene encodes the imidazole acetol phosphate transaminase enzyme (E.C.2.6.1.9), which is responsible for converting imidazole acetol-P to histidinol-P in step 8 of histidine biosynthesis. The nucleotide sequences of a 2196 by gene fragment and a corresponding cDNA clone were determined. Three intron sequences punctuate the 1451 by coding region which generates a predicted polypeptide of 384 amino acids with a molecular mass of 42736 daltons. Northern analysis of his3 mRNAs indicates that the transcript is approximately 1.6 kb in size. Steady-state levels are down-regulated by nitrogen limitation but are unaffected by histidine starvation. The deduced amino acid sequence was compared to the Saccharomyces cerevisiae HIS5, Escherichia coli HisC, and Salmonella typhimurium HisC proteins, all of which are imidazole acetol phosphate transaminases. The S. pombe his3 protein was 49.5% identical to the S. cerevisiae HISS protein and 21.5% identity was found when all four proteins were compared. The shuttle vector pBGI was constructed by subcloning the smallest functional region of his3 and the S. pombe arsl sequence into pUC18 for use in transformation of His3 — S. pombe strains. New S. pombe strains in which the his3 gene was deleted have also been constructed.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00391010
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