Domain structure of a mammalian myosin I beta.
We have determined the primary structure of a myosin I (called mammalian myosin I beta, MMI beta) from bovine brain and identified its functional domains. The protein was previously purified from brain and adrenal gland. Several constructs were generated and expressed in Escherichia coli as glutathione S-transferase fusion proteins and the recombinant proteins were recognized by monoclonal antibodies that recognize either "head" or "tail" domains of native myosin I. A gel overlay method was used to confirm that calmodulin binds to the consensus calmodulin-binding sequence in MMI beta. Binding assays were used to detect interaction with anionic phospholipid vesicles. We conclude that MMI beta consists of an amino-terminal 80.5-kDa domain that contains the ATP- and actin-binding sites, followed by an 8.5-kDa domain with three calmodulin-binding sequences and a basic 30-kDa carboxyl-terminal tail segment that binds to anionic phospholipids and membranes.
Reizes
O
O
Department of Pharmacology, University of Texas-Southwestern Medical Center at Dallas 75235.
Barylko
B
B
Li
C
C
Südhof
T C
TC
Albanesi
J P
JP
D000199:Actins / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001923:Brain Chemistry; D002147:Calmodulin / Q000378:metabolism; D002417:Cattle; D002645:Chickens; D003001:Cloning, Molecular; D016384:Consensus Sequence; D017931:DNA Primers; D004926:Escherichia coli; D005982:Glutathione Transferase / Q000096:biosynthesis; D008322:Mammals / Q000378:metabolism*; D008969:Molecular Sequence Data; D009218:Myosins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D000199:Actins; D002147:Calmodulin; D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D005982:Glutathione Transferase; D009218:Myosins