CHARACTERIZATION OF 2 PUTATIVE PROTEIN SER THR KINASES FROM ACTINOMYCETE STREPTOMYCES GRANATICOLOR BOTH ENDOWED WITH DIFFERENT PROPERTIES/

The structural genes, pkg4 and pkg3, encoding two putative protein ser ine/threonine kinases in Streptomyces granaticolor; have been cloned a nd sequenced. The genes were isolated after screening genomic sublibra ries with specific probes obtained by PCR amplification of chromosomal DNA using: degenerate primers which correspond to amino acid sequence s highly conserved in eukaryotic protein Ser/Thr kinases. The sequence s of these genes predict polypeptide chains of 761 and 780 amino acids for Pkg4 and Pkg3, respectively. The genes are separated by only 2 bp and therefore probably constitute an operon. pkg4, which is positione d upstream of pkg3, contains a UUA(Leu) codon suggesting a development al-dependent mode of expression. The amino-terminal half of both prote ins clearly shares similarities with the family of protein Ser/Thr kin ases. Both proteins studied also possess a region rich in Pro and Ala residues and a repeating motif of 11 amino acid residues, the function of which is unknown, in the carboxy-terminal domain. Expression of pk g4 in Escherichia coli gave rise to two different forms: a soluble pro tein autophosphorylated at threonine residues and an insoluble form ph osphorylated at threonine and serine residues. In contrast, when pkg3 was expressed in E. coli, no autophosphorylation was detected either i n vivo or in vitro.