Myosin regulatory light chain phosphorylation by conventional mvosin light chain kinase(MLCK) plays a key role in the initiation of smooth muscle con traction and many cortractile motile processes in monuscle cells. Previous studies showed conventional MLCK binds tightly to actio-myosin containing filaments. The N-terminal 661 residues of MICK but no C terminal telokin fragment are essential for hgh affinity binding to smooth muscle myofilaments and fibroblast strees fbers (Lin et al. JBC 272;7412, 1997). Additionaly the apparent affinity was greate for myofilaments compared to F actia. To define this binding domain more precisely, a senes of MLCK N-terminal deletion mutants and N-terminal GST fusion proteins were constructed and espressed either in COS-7 cells or in E coli. Binding to smooth muscle myofilaments was measured by cosedimentation assvise followed by Western blotting. Results show the N-terminal 75 amino smooth muscle myofilaments as fightly as the following kinase. Results aso show that the core binding sequetnce of this segment is between residues 23 and 40, The primary sequence from residues 23 to 10 shows no sequence similarity to other known protein binding motifs by comparison to sequences in the protein data base. Therefore, this myofilament binding sequence may represent a novel protein binding motif.(supported by Bashor Research Fund and NIH Grat HL26043).
|Original language||English (US)|
|State||Published - Dec 1 1997|
ASJC Scopus subject areas
- Molecular Biology