Molecular Cloning and Characterization of a Novel Mammalian Protein Kinase Harboring a Homology Domain that Defines a Subfamily of Serine/Threonine Kinases

Authors


  • Note. The novel nucleotide sequence data published here have been deposited with the EMBI sequence data bank and are available under the accession number X89383

H-G. Joost, Institut für Pharmakologie und Toxikologie, Medizinische Fakultät der RWTH Aachen, Wendlingweg 2, D-52057 Aachen, Germany
Fax: +49 8888433.

Abstract

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 approach based on the polymerase chain reaction. The encoded polypeptide (746 amino acids, Mr= 81627) contains all conserved subdomains characteristic of the protein serine/threonine kinase family. A recombinant fusion protein with glutathione S -transferase catalysed autophosphorylation 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-LI cells, SNRK was specifically expressed in the differentiated, adipocyte-like phenotype, where as its mRNA was not detected in fibroblasts. Sequence comparisons of its catalytic 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 consecutive acidic residues. This part of the sequence exhibits no extended similarity with other protein In addition, we detected a high degree of sequence similarity with other SNF1 -related protein kinases in a small region (30–35 amino acids) flanking the C-terminus of the catalytic domain. This domain (designated the SNH domain) appears to define the subfamily of SNF1-related protein kinases and might represent a new type of regulatory domain of protein kinases.

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