Molecular charge dominates the inhibition of actomyosin in skinned muscle fibers by SH1 peptides

• Published in: Biophysical journal 1991-08, Vol.60 (2), p.352-359
• Main Authors: Chase, P.B., Beck, T.W., Bursell, J., Kushmerick, M.J.
• Format: Article
• Language: English
• Subjects: Actomyosin - antagonists & inhibitors; Actomyosin - chemistry; Actomyosin - metabolism; Amino Acid Sequence; Animals; Binding Sites; Biological and medical sciences; Biophysical Phenomena; Biophysics; Cell physiology; Electrochemistry; Fundamental and applied biological sciences. Psychology; In Vitro Techniques; Molecular and cellular biology; Molecular Sequence Data; Muscle Contraction - physiology; Muscles - metabolism; Myosin Subfragments - chemistry; Myosin Subfragments - metabolism; Myosin Subfragments - pharmacology; Rabbits
• ISSN: 0006-3495; 1542-0086
• DOI: 10.1016/S0006-3495(91)82060-4

It is not definitively known whether the highly conserved region of myosin heavy chain around SH1 (Cys 707) is part of the actin-binding site. We tested this possibility by assaying for competitive inhibition of maximum Ca-activated force production of skinned muscle fibers by synthetic peptides which had sequences derived from the SH1 region of myosin. Force was inhibited by a heptapeptide (IRICRKG) with an apparent K0.5 of about 4 mM. Unloaded shortening velocity of fibers, determined by the slack test, and maximum Ca-activated myofibrillar MgATPase activity were also inhibited by this peptide, but both required higher concentrations. We found that other cationic peptides also inhibited force in a manner that depended on the charge of the peptide; increasing the net positive charge of the peptide increased its efficacy. The inhibition was not significantly affected by altering solution ionic strength (100–200 mM). Disulfide bond formation was not involved in the inhibitory mechanism because a peptide with Thr substituted for Cys was inhibitory in the presence or absence of DTT. Our data demonstrate that the net charge was the predominant molecular characteristic correlated with the ability of peptides from this region of myosin heavy chain to inhibit force production. Thus, the hypothesis that the SH1 region of myosin is an essential part of the force-producing interaction with actin during the cross-bridge cycle (Eto, M., R. Suzuki, F. Morita, H. Kuwayama, N. Nishi, and S. Tokura., 1990, J. Biochem. 108:499–504; Keane et al., 1990, Nature (Lond.). 344:265–268) is not supported.