The C-terminal helix in subdomain 4 of the regulatory light chain is essential for myosin regulation.
In vertebrate smooth/non-muscle myosins, phosphorylation of the regulatory light chains by a specific calmodulin-activated kinase controls both myosin head interaction with actin and assembly of the myosin into filaments. Previous studies have shown that the C-terminal domain of the regulatory light chain is crucial for the regulation of these myosin functions. To further dissect the role of this region of the light chain in myosin regulation, a series of chicken smooth muscle myosin regulatory light chain mutants has been constructed with successive C-terminal deletions. These mutants were synthesized in Escherichia coli and analysed by their ability to restore Ca2+ regulation to scallop myosin that had been stripped of its native regulatory light chains ('desensitized'). The results show that regulatory light chain mutants with deletions in the C-terminal helix in subdomain 4 were able to reform the regulatory Ca2+ binding site on the scallop myosin head, but had lost the ability to suppress scallop myosin filament assembly and interaction with actin in the absence of Ca2+. Further deletions in the C-terminal domain led to a gradual loss of ability to restore the regulatory Ca2+ binding site. Thus, the regions in the C-terminal half of the regulatory light chain responsible for myosin regulation can be identified.
Rowe
T
T
MRC Laboratory of Molecular Biology, Cambridge, UK.
Kendrick-Jones
J
J
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism; D002645:Chickens; D003001:Cloning, Molecular; D004247:DNA; D004926:Escherichia coli; D008969:Molecular Sequence Data; D008974:Mollusca; D009130:Muscle, Smooth / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009218:Myosins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D012995:Solubility
D000199:Actins; D004247:DNA; D009218:Myosins; D002118:Calcium