Incorporation of an azobenzene derivative into the energy transducing site of skeletal muscle myosin results in photo-induced conformational changes.
The region containing reactive cysteines, Cys 707 (SH1)-Cys 697 (SH2), of skeletal muscle myosin is thought to play a key role in the conformational changes of the myosin head during force generation coupled to ATP hydrolysis. In the present study, we synthesized a photochromic crosslinker, 4,4'-azobenzene-dimaleimide (ABDM), that undergoes reversible cis-trans isomerization upon ultra violet (UV) and visible (VIS) light irradiation resulting in a change in the crosslinking length from 5 to 17 A. The reactive cysteines, SH1 and SH2, of myosin subfragment 1 (S1) were crosslinked with ABDM, yielding an ABDM-S1 complex. The changes in absorbance induced by UV/VIS light irradiation of the complex were similar to those of free ABDM indicating that the incorporation of ABDM at the SH1 and SH2 sites did not disrupt the isomerization of crosslinked ABDM. Small-angle synchrotron X-ray scattering analysis of the ABDM-S1 complex in solution suggested that the localized conformational changes resulting from the cis to trans isomerization on ABDM crosslinking of SH1 and SH2 induced a small but significant swing in the lever arm portion of S1 in the opposite direction from that induced by ATP.
Umeki
Nobuhisa
N
Division of Bioengineering, Graduate School of Engineering, Soka University, Hachioji, Tokyo 192-8577, Japan.
Yoshizawa
Tsuyoshi
T
Sugimoto
Yasunobu
Y
Mitsui
Toshiaki
T
Wakabayashi
Katsuzo
K
Maruta
Shinsaku
S
D000818:Animals; D001391:Azo Compounds / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D001940:Breast / Q000378:metabolism; D002645:Chickens; D003432:Cross-Linking Reagents; D003545:Cysteine / Q000378:metabolism; D004734:Energy Metabolism; D008027:Light; D015394:Molecular Structure; D015879:Myosin Subfragments / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D024744:Skeletal Muscle Myosins / Q000378:metabolism*
D001391:Azo Compounds; D003432:Cross-Linking Reagents; D015879:Myosin Subfragments; D024744:Skeletal Muscle Myosins; C009850:azobenzene; D003545:Cysteine