title: Modular structure of smooth muscle Myosin light chain kinase: hydrodynamic modeling and functional implications.
abstract: Smooth muscle myosin light chain kinase (smMLCK) is a calcium-calmodulin complex-dependent enzyme that activates contraction of smooth muscle. The polypeptide chain of rabbit uterine smMLCK (Swiss-Prot entry P29294) contains the catalytic/regulatory domain, three immunoglobulin-related motifs (Ig), one fibronectin-related motif (Fn3), a repetitive, proline-rich segment (PEVK), and, at the N-terminus, a unique F-actin-binding domain. We have evaluated the spatial arrangement of these domains in a recombinant 125 kDa full-length smMLCK and its two catalytically active C-terminal fragments (77 kDa, residues 461-1147, and 61 kDa, residues 461-1002). Electron microscopic images of smMLCK cross-linked to F-actin show particles at variable distances (11-55 nm) from the filament, suggesting that a well-structured C-terminal segment of smMLCK is connected to the actin-binding domain by a long, flexible tether. We have used structural homology and molecular dynamics methods to construct various all-atom representation models of smMLCK and its two fragments. The theoretical sedimentation coefficients computed with HYDROPRO were compared with those determined by sedimentation velocity. We found agreement between the predicted and observed sedimentation coefficients for models in which the independently folded catalytic domain, Fn3, and Ig domains are aligned consecutively on the long axis of the molecule. The PEVK segment is modeled as an extensible linker that enables smMLCK to remain bound to F-actin and simultaneously activate the myosin heads of adjacent myosin filaments at a distance of >or=40 nm. The structural properties of smMLCK may contribute to the elasticity of smooth muscle cells.
authors.affiliation: Boston Biomedical Research Institute, Watertown, Massachusetts 02472-2829, USA.
authors.forename: Yasuko
authors.initials: Y
authors.lastname: Mabuchi
authors.forename: Katsuhide
authors.initials: K
authors.lastname: Mabuchi
authors.forename: Walter F
authors.initials: WF
authors.lastname: Stafford
authors.forename: Zenon
authors.initials: Z
authors.lastname: Grabarek
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D004548:Elasticity; D005260:Female; D008854:Microscopy, Electron; D009130:Muscle, Smooth / Q000201:enzymology*; D009219:Myosin-Light-Chain Kinase / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D014599:Uterus
chemical_list: D000199:Actins; D009219:Myosin-Light-Chain Kinase