Size and Charge Requirements for Kinetic Modulation and Actin Binding by Alkali 1-type Myosin Essential Light Chains

David J. Timson, Hylary R. Trayer, K. John Smith, Ian P. Trayer
1999 Journal of Biological Chemistry  
The alkali 1-type isoforms of myosin essential light chains from vertebrate striated muscles have an additional 40 or so amino acids at their N terminus compared with the alkali 2-type. Consequently two light chain isoenzymes of myosin subfragment-1 can be isolated. Using synthesized peptide mimics of the N-terminal region of alkali 1-type essential light chains, we have found by 1 H NMR that the major actin binding region occurred in the N-terminal four residues, APKK. . . . These results were
more » ... These results were confirmed by mutating this region of the human atrial essential light chain, resulting in altered actin-activated MgATPase kinetics when the recombinant light chains were hybridized into rabbit skeletal subfragment 1. Substitution of either Lys 3 or Lys 4 with Ala resulted in increased K m and k cat and decreased actin binding (as judged by chemical cross-linking). Replacement of Lys 4 with Asp reduced actin binding and increased K m and k cat still further. Alteration of Ala 1 to Val did not alter the kinetic parameters of the hybrid subfragment 1 or the essential light chain's ability to bind actin. Furthermore, we found a significant correlation between the apparent K m for actin and the k cat for MgATP turnover for each mutant hybrid, strengthening our belief that the binding of actin by alkali 1-type essential light chains results directly in modulation of the myosin motor.
doi:10.1074/jbc.274.26.18271 pmid:10373429 fatcat:i3ef22ajfrbi5lc6ou6cxwu2ke