We report here the cloning and characterization of the gene encoding the 130-kDa myosin heavy chain kinase (MHCK A) from the amoeba Dictyostelium. Previous studies have shown that purified MHCK A phosphorylates threonines in the carboxyl-terminal tail portion of the Dictyostelium myosin II heavy chain and that phosphorylation of these sites is critical in regulating the assembly and disassembly of myosin II filaments in vitro and in vivo. Biochemical analysis of MHCK A, together with analysis of the primary sequence, suggests that the amino-terminal 500 amino acids form an -helical coiled-coil domain and that residues from position 860 to the carboxyl terminus (residue 1146) form a domain with significant similarity to the -subunit of heterotrimeric G proteins. No part of the MHCK A sequence displays significant similarity to the catalytic domain of conventional eukaryotic protein kinases. However, both native and recombinant MHCK A displayed autophosphorylation activity following renaturation from SDS gels, and MHCK A expressed in Escherichia coli phosphorylated purified Dictyostelium myosin, confirming that MHCK A is a bona fide protein kinase. Cross-linking studies demonstrated that native MHCK A is a multimer, consistent with the presence of an amino-terminal coiled-coil domain. Southern blot analysis indicates that MHCK A is encoded by a single gene that has no detectable introns.

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