Modulation of cross-bridge affinity for MgGTP by Ca2+ in skinned fibers of rabbit psoas muscle.
Previously we reported that saturation of cross-bridges with MgATP gamma S in skinned muscle fibers was calcium sensitive. In the present study we investigate whether this observation can be generalized to other nucleotides by studying saturation of cross-bridges with MgGTP. In solution, myosin-subfragment 1 (S1) in the presence of 10 mM MgGTP was found to bind to actin with low affinity, similar to that in the presence of MgATP and MgATP gamma S. In EGTA buffer, the equatorial x-ray diffraction intensity ratio I11/I10 recorded in single skinned fibers decreased upon increasing MgGTP concentration from 0 to 10 mM (1 degree C and 170 mM ionic strength). The I11/I10 ratio leveled off at 10 mM MgGTP, indicating full saturation of cross-bridges with the nucleotide. Under these conditions, the value of I11/I10 is indistinguishable from that obtained in the presence of saturating [MgATP]. In CaEGTA buffer, however, the decrease in I11/I10 occurs over a wider range of concentrations, and there is no indication of I11/I10 leveling off at 10 mM MgGTP, suggesting that full saturation is not reached. The Ca2+ dependence of GTP binding appears to be a direct consequence of the differences in the affinities of the strongly bound cross-bridges to actin versus weakly bound cross-bridges to actin. A biochemical scheme that could qualitatively explain the titration behavior of ATP gamma S and GTP is presented.
Frisbie
S M
SM
National Institute of Arthritis, Musculoskeletal, and Skin Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Chalovich
J M
JM
Brenner
B
B
Yu
L C
LC
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D007700:Kinetics; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D009210:Myofibrils; D016658:Psoas Muscles / Q000378:metabolism; D011817:Rabbits; D014961:X-Ray Diffraction
C036852:magnesium GTP; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D002118:Calcium