Autophosphorylation of smooth muscle myosin light chain kinase at its regulatory domain.
Autophosphorylation of smooth muscle myosin light chain kinase was initially reported by Foyt et al. [Foyt, H. L., & Means, A. R. (1985) J. Cyclic Nucleotide Protein Phosphorylation Res. 260, 8978-8983], however, the effects of autophosphorylation on the kinase activity as well as the location of the sites have not been elucidated. Here we demonstrate that MLCK is autophosphorylated at three sites, Thr 803, Ser 815, and Ser 823, and this phosphorylation alters MLCK activity. Two phosphorylation sites are located in the regulatory domain of the kinase, the threonine site toward the autoinhibitory region and the serine site (Ser 815) in close proximity to the calmodulin anchoring site. The autophosphorylation was significantly inhibited by the binding of calmodulin. The autophosphorylation at Thr 803 is an intramolecular process, and the alignment of the basic amino acid residues nearby Thr 803 was highly homologous to the phosphorylation site of myosin light chain, suggesting that the regulatory site is in close proximity to the catalytic site in the three-dimensional structure. The phosphorylation at the threonine site activated the calmodulin-independent activity while the phosphorylation at the serine site inhibited the calmodulin-dependent activity due to a decrease in the affinity for calmodulin. This finding shows another example of the activation of calmodulin-dependent kinases by autophosphorylation at its autoinhibitory region and provides a new clue for understanding the calmodulin/MLCK signalling pathway.
Tokui
T
T
Department of Physiology and Biophysics, Case Western Reserve University, School of Medicine, Cleveland, Ohio 44106, USA.
Ando
S
S
Ikebe
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D004789:Enzyme Activation; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009130:Muscle, Smooth / Q000201:enzymology*; D009219:Myosin-Light-Chain Kinase / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D017386:Sequence Homology, Amino Acid; D014422:Turkeys
D009219:Myosin-Light-Chain Kinase