STRUCTURE AND FUNCTION OF THE 10S CONFORMATION OF SMOOTH-MUSCLE MYOSIN

Smooth myosin regulatory light chain (RLC) was exchanged with RLC labe led with benzophenone-4-iodoacetamide at Cys-108. Irradiation under co nditions that favor the folded (10 S) conformation resulted in 10 S cr oss-linked myosin that could not unfold. Purified 10 S cross-linked my osin was cross-linked between the RLC of one head to light meromyosin between leucine 1554 and glutamate 1583, adjacent to a predicted nonco iled region, approximately 60 nn from the tip of the tail. At high ion ic strength without actin, product release from one-half of the heads was slow (like 10 S) whereas the other half were activated. This sugge sts that tail binding to the RLC carboxyl-terminal domain stabilizes i onic interactions important to slow nucleotide release. With actin, pr oduct release from both (un)phosphorylated 10 S cross-linked myosin wa s from one slow population similar to unphosphorylated filaments. 10 S cross-linked myosin weakly bound actin (dissociation constant > 500 m u M) and did not move actin in vitro. Single-headed myosin did not fol d or trap nucleotide, These and other data suggest that ''trapping'' o ccurs only with both heads and the tail binds to a newly formed site, which includes the RLC carboxyl-terminal domain, once trapping has occ urred.