Cloning and sequence analysis of the glyceraldehyde-3-phosphate dehydrogenase gene from the zygomycetes fungus Rhizomucor miehei

摘要

Rhizomucor miehei is important from a biotechnological aspect in consequence of its content of aspartic proteinase, which has high milk-clotting activity. A genomic library of R. miehei NRRL 5901 has been constructed in a phage (Lambda Fix II) vector. The glyceraldehyde-3-phosphate dehydrogenase (gpd) gene was isolated from this library by hybridization of the recombinant phage clones with a gpd-specific gene probe generated by polymerase chain reaction. The complete nucleotide sequence encodes a putative polypeptide chain of 336 amino acids interrupted by 5 introns. The predicted amino acid sequence of this gene shows a high degree of sequence similarity to the glyceraldehyde-3-phosphate dehydrogenase proteins from yeast and filamentous fungi. The promoter region, containing a consensus TATA box, and 246-bp downstream from the putative stop codon were also determined. The possibility of using the gpd promoter in the construction of new transformation vectors is discussed.