STREPTOMYCES ATP NUCLEOTIDE 3'-PYROPHOSPHOKINASE-GENE CLONING AND SEQUENCE-ANALYSIS

Streptomyces ATP nucleotide 3'-pyrophosphokinase is an extracellular e nzyme that transfers 5'-beta,gamma-pyrophosphoryl groups of ATP to a v ariety of nucleotides at the 3'-OH site. The enzyme gene was cloned fr om partially Sau3AI-digested chromosomal DNA of S. morookaensis in S. lividans TK24/pIJ699 and then in E. coli JM83/pUC12. Some transformant s produced the active enzyme. The gene was sequenced by the dideoxynuc leotide termination procedure, Its GC content was 72%. Its putative pr omoter regions, shoeing little homology to that of the Streptomyces co nsensus type, were pointed out. No sequence homology was found between the pyrophosphokinase and any other known genes including those of th e most mechanistically similar bacterial stringent factor and related proteins. Northern hybridization analysis showed that the gene is cons titutionally polycistronic and expressed under transcriptional control . Nuclease S1 mapping indicated that the gene transcription starts fro m its translation initiation site.