Kinetic analysis of Drosophila muscle myosin isoforms suggests a novel mode of mechanochemical coupling.
The molecular mechanism of myosin function was addressed by measuring transient kinetic parameters of naturally occurring and chimeric Drosophila muscle myosin isoforms. We assessed the native embryonic isoform, the native indirect flight muscle isoform, and two chimeric isoforms containing converter domains exchanged between the indirect flight muscle and embryonic isoforms. Myosin was purified from the indirect flight muscles of transgenic flies, and S1 was produced by alpha-chymotryptic digestion. Previous studies in vertebrate and scallop myosins have shown a correlation between actin filament velocity in motility assays and cross-bridge detachment rate, specifically the rate of ADP release. In contrast, our study showed no correlation between the detachment rate and actin filament velocity in Drosophila myosin isoforms and further that the converter domain does not significantly influence the biochemical kinetics governing the detachment of myosin from actin. We suggest that evolutionary pressure on a single muscle myosin gene may maintain a fast detachment rate in all isoforms. As a result, the attachment rate and completion of the power stroke or the equilibrium between actin.myosin.ADP states may define actin filament velocity for these myosin isoforms.
D000199:Actins / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D017301:Ca(2+) Mg(2+)-ATPase / Q000737:chemistry; D002918:Chymotrypsin / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004331:Drosophila melanogaster / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008027:Light; D008274:Magnesium / Q000737:chemistry; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000378:metabolism; D009132:Muscles / Q000378:metabolism*; D009218:Myosins / Q000737:chemistry* / Q000378:metabolism; D010782:Photolysis; D011485:Protein Binding; D020033:Protein Isoforms; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011817:Rabbits; D012542:Scattering, Radiation; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
D000199:Actins; D020033:Protein Isoforms; D000255:Adenosine Triphosphate; D002918:Chymotrypsin; C117608:alpha-chymotrypsin; D000251:Adenosine Triphosphatases; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins; D008274:Magnesium