TY - JOUR
T1 - Substitution mutations in the myosin essential light chain lead to reduced actin-activated ATPase activity despite stoichiometric binding to the heavy chain
AU - Ho, Guyu
AU - Chisholm, Rex L.
PY - 1997
Y1 - 1997
N2 - Myosin essential light chain (ELC) wraps around an α-helix that extends from the myosin head, where it is believed to play a structural support role. To identify other role(s) of the ELC in myosin function, we have used an alanine scanning mutagenesis approach to convert charged residues in loops I, H, HI, and helix G of the Dictyostelium ELC into uncharged alanines. Dictyostelium was used as a host system to study the phenotypic and biochemical consequences associated with the mutations. The ELC carrying loop mutations bound with normal stoichiometry to the myosin heavy chain when expressed in ELC-minus cells. When expressed in wild type cells these mutants competed efficiently with the endogenous ELC for binding, suggesting that the affinity of their interaction with the heavy chain is comparable to that of wild type. However, despite apparently normal association of ELC the cells still exhibited a reduced efficiency to undergo cytokinesis in suspension. Myosin purified from these cells exhibited 45-fold reduction in actin- activated ATPase activity and a decrease in motor function as assessed by an in vitro motility assay. These results suggest that the ELC contributes to myosin's enzymatic activity in addition to providing structural support for the α-helical neck region of myosin heavy chain.
AB - Myosin essential light chain (ELC) wraps around an α-helix that extends from the myosin head, where it is believed to play a structural support role. To identify other role(s) of the ELC in myosin function, we have used an alanine scanning mutagenesis approach to convert charged residues in loops I, H, HI, and helix G of the Dictyostelium ELC into uncharged alanines. Dictyostelium was used as a host system to study the phenotypic and biochemical consequences associated with the mutations. The ELC carrying loop mutations bound with normal stoichiometry to the myosin heavy chain when expressed in ELC-minus cells. When expressed in wild type cells these mutants competed efficiently with the endogenous ELC for binding, suggesting that the affinity of their interaction with the heavy chain is comparable to that of wild type. However, despite apparently normal association of ELC the cells still exhibited a reduced efficiency to undergo cytokinesis in suspension. Myosin purified from these cells exhibited 45-fold reduction in actin- activated ATPase activity and a decrease in motor function as assessed by an in vitro motility assay. These results suggest that the ELC contributes to myosin's enzymatic activity in addition to providing structural support for the α-helical neck region of myosin heavy chain.
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U2 - 10.1074/jbc.272.7.4522
DO - 10.1074/jbc.272.7.4522
M3 - Article
C2 - 9020178
AN - SCOPUS:0031055280
SN - 0021-9258
VL - 272
SP - 4522
EP - 4527
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 7
ER -