Direct chemical evidence that serine 180 in the glycine-rich loop of myosin binds to ATP.
The photochemical reaction of MgADP-vanadate with the active site of myosin has been used to place a serine at the binding site for the gamma-phosphate of ATP. Irradiation of the MgADP-vanadate myosin subfragment 1 transition state-like complex with UV light specifically photooxidizes the hydroxyl group of a serine residue to an aldehyde (Cremo, C. R., Grammer, J. C., and Yount, R. G. (1988) Biochemistry 27, 8415-8420). Reduction of photooxidized myosin with Na-B3H4 gave only 3H-labeled serine. Here, subsequent extensive proteolytic digestion of 3H-labeled myosin subfragment 1 with trypsin and thermolysin yielded two 3H-labeled peptides, both of which contained the sequence Gly-Glu-Ser-Gly-Ala-Gly-Lys-Thr, in which all the 3H was associated with the serine. This sequence is conserved in all myosin heavy chains sequenced to date and corresponds to residues 178-185 in the rabbit myosin heavy chain (Tong, S. W., and Elzinga, M. (1983) J. Biol. Chem. 21, 13100-13110). These results place Ser-180 at the gamma-phosphate-binding site for ATP and indicate that the glycine-rich loop around the serine provides essential elements of the phosphate-binding site for ATP in all myosin molecules. Such a role was previously suggested based on the common sequence Gly-X-X-X-X-Gly-Lys-Thr/Ser, found in myosin and many other nucleotide-binding enzymes (Walker, J. E., Saraste, M., Runswick, M. H., and Gay, N. J. (1982) EMBO J. 1, 945-951).
Cremo
C R
CR
Biochemistry/Biophysics Program, Washington State University Pullman 99164-4660.
Grammer
J C
JC
Yount
R G
RG
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D009132:Muscles / Q000201:enzymology*; D009218:Myosins / Q000378:metabolism*; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000032:analysis; D010777:Photochemistry; D011817:Rabbits; D012694:Serine; D014639:Vanadium
D010446:Peptide Fragments; D014639:Vanadium; D012694:Serine; D000255:Adenosine Triphosphate; D009218:Myosins