Mutational analysis of an actin-binding site of cofilin and characterization of chimeric proteins between cofilin and destrin.
Cofilin and destrin are two related low molecular weight mammalian actin-binding proteins. Cofilin is an F-actin side-binding and pH-dependent actin-depolymerizing protein, and destrin is a pH-independent actin-depolymerizing protein. We have introduced a few point mutations within an actin-binding sequence of cofilin. Biochemical analyses of these mutant proteins have clearly shown that Lys112 and Lys114 of cofilin are crucially but differently involved in its interaction with actin and phosphatidylinositol 4,5-bisphosphate. This is the first example among actin-binding proteins whose point mutations inactivate their interaction with actin in vitro. We have also made and characterized a series of chimeric proteins between cofilin and destrin to identify the regions responsible for the pH dependence and the F-actin side binding activity of cofilin. Our results suggest that a central region consisting of 42 amino acid residues and a carboxyl-terminal quarter of cofilin are both involved in regulation of the pH-dependent actin depolymerizing activity and the activity to bind along F-actin.
Moriyama
K
K
Department of Biophysics and Biochemistry, Faculty of Science, University of Tokyo, Japan.
Yonezawa
N
N
Sakai
H
H
Yahara
I
I
Nishida
E
E
D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins; D051345:Destrin; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008840:Microfilament Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D051339:Actin Depolymerizing Factors; D000199:Actins; D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D051345:Destrin; D008840:Microfilament Proteins; D009419:Nerve Tissue Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D011993:Recombinant Fusion Proteins