Design and functional analysis of actomyosin motor domain chimera proteins.
To gain more structural and functional information on the actomyosin complexes, we have engineered chimera proteins carrying the entire Dictyostelium actin in the loop 2 sequence of the motor domain of Dictyostelium myosin II. Although the chimera proteins were unable to polymerize by themselves, addition of skeletal actin promoted polymerization. Electron microscopic observation demonstrated that the chimera proteins were incorporated into actin filaments, when copolymerized with skeletal actin. Copolymerization with skeletal actin greatly enhanced the MgATPase, while the chimera proteins without added skeletal actin hydrolyzed ATP at a very low rate. These results indicate that the actin part and the motor domain part of the chimera proteins are correctly folded, but the chimera proteins are structurally stressed so that efficient polymerization is inhibited.
Yokoyama
Keiichi
K
Department of Metallurgy, Graduate School of Engineering, Tohoku University, Aramaki-aza-Aoba 02, Aobaku, Sendai 980-8579, Japan.
Hiratuka
Yuichi
Y
Akimaru
Erika
E
Hirose
Keiko
K
Uyeda
Taro Q P
TQ
Suzuki
Makoto
M
D008841:Actin Cytoskeleton / Q000648:ultrastructure; D000199:Actins / Q000235:genetics*; D000205:Actomyosin / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002645:Chickens; D004023:Dictyostelium; D020409:Molecular Motor Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D008969:Molecular Sequence Data; D024462:Myosin Type II / Q000235:genetics*; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000302:isolation & purification / Q000648:ultrastructure; D016415:Sequence Alignment
D000199:Actins; D020409:Molecular Motor Proteins; D011993:Recombinant Fusion Proteins; D000205:Actomyosin; D000251:Adenosine Triphosphatases; D024462:Myosin Type II