A new Acanthamoeba myosin heavy chain. Cloning of the gene and immunological identification of the polypeptide.
An Acanthamoeba myosin heavy chain has been identified whose tail domain amino acid sequence distinguishes it from Acanthamoeba myosins IB, IC, and II. The gene for this novel myosin heavy chain spans approximately 6.8 kilobases, is split by 17 introns, and encodes a 177-kDa polypeptide. While the amino-terminal approximately 90 kDa of this polypeptide is highly similar to the globular head sequences of myosins I and II, its approximately 87-kDa tail domain shows essentially no similarity to the tail sequences of either type of myosin. The only exception to this is the carboxyl-terminal approximately 50-amino acid region of the polypeptide, which is homologous to the carboxyl termini of the myosins I. Interestingly, this approximately 50-residue segment has been shown to exist in a diverse family of cytoskeleton-associated proteins that include nonreceptor tyrosine kinases, phospholipase C gamma, and fodrin (Rodaway, A. R. F., Sternberg, M. J. E., and Bentley, D. L. (1989) Nature 342, 624). Sequence analysis indicates that the tail domain of this new myosin is incapable of forming a myosin II-like coiled-coil structure, implying that the protein is single-headed and nonfilamentous. For this reason we have tentatively classified it as a high molecular weight form of myosin I (HMWMI). To determine if HMWMI exists in cells, antiserum was raised against a bacterially expressed fusion peptide made using a cDNA clone encoding most of the unique HMWMI tail domain. This antiserum does not recognize Acanthamoeba myosins IB, IC, or II but does recognize a single polypeptide in whole cell extracts with the mobility predicted for the HMWMI heavy chain. This protein is precipitated from crude extracts using F-actin and released from the pellet by ATP, supporting its classification as a member of the myosin family of proteins.
Horowitz
J A
JA
Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
Hammer
J A
JA
D000048:Acanthamoeba / Q000235:genetics* / Q000378:metabolism; D000205:Actomyosin / Q000302:isolation & purification / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D015723:Gene Library; D005796:Genes; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009218:Myosins / Q000235:genetics* / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid
D046911:Macromolecular Substances; D011993:Recombinant Fusion Proteins; D000205:Actomyosin; D009218:Myosins