MOLECULAR-CLONING AND CHARACTERIZATION OF A NOVEL MAMMALIAN PROTEIN-KINASE HARBORING A HOMOLOGY DOMAIN THAT DEFINES A SUBFAMILY OF SERINE THREONINE KINASES

The cDNA of a novel protein kinase (referred to as SNRK) was isolated from a rat fat cell cDNA library with a probe generated by a cloning a pproach based on the polymerase chain reaction. The encoded polypeptid e (746 amino acids, M(r) = 81627) contains all conserved subdomains ch aracteristic of the protein serine/threonine kinase family. A recombin ant fusion protein with glutathione S-transferase catalysed autophosph orylation as well as phosphorylation of histone, confirming that SNRK has indeed protein kinase activity, By Northern blot hybridization, a 5-kb mRNA was detected in brain, heart, fat cells, intestine, testis, ovary, adrenal gland and thymus. In 3T3-L1 cells, SNRK was specificall y expressed in the differentiated, adipocyte-like phenotype, whereas i ts mRNA was not detected in fibroblasts. Sequence comparisons of its c atalytic domain relate SNRK to the SNF1 family of protein kinases. The noncatalytic domain comprises several intriguing structural features, including a glycine-rich region, two PEST sequences, and a bipartite nuclear localization signal which is preceded by a stretch of ten cons ecutive acidic residues. This part of the sequence exhibits no extende d similarity with other proteins, In addition, we detected a high degr ee of sequence similarity with other SNF1-related protein kinases in a small region (30-35 amino acids) flanking the C-terminus of the catal ytic domain. This domain (designated the SNH domain) appears to define the subfamily of SNF1-related protein kinases and might represent a n ew type of regulatory domain of protein kinases.