Detection of a sequence involved in actin-binding and phosphoinositide-binding in the N-terminal side of cofilin.
Cofilin is an actin-binding protein of low molecular weight which is widely distributed in eukaryotes and is deeply involved in the dynamics of actin assembly in the cytoplasm. The actin-binding ability of cofilin is inhibited by inositol phosphates (PIP2), and the PIP2- and actin-binding site(s) has been localized in residues W104-M115 of the cofilin primary sequence (Yonezawa et al. 1991 ). In the present study, in order to further clarify the functional domains in cofilin molecule, we constructed expression vectors containing cDNAs of different size with deletion at the 3'-region of the open reading frame. The truncated cofilin molecules produced in E. coli were purified and examined for their actin-binding and PIP2-binding ability. We found that the truncated cofilin molecule without C-terminal residues #100-#166 including the previously-described actin-binding site could be cross-linked with actin by EDC, a zero-length cross-linker. In addition, these truncated peptides as well as synthetic peptides corresponding to the N-terminal sequence of cofilin suppressed the inhibitory action of PIP2 on actin-cofilin interaction. These results strongly suggest that additional actin- and PIP2-binding sites exist in the N-terminal region of cofilin.
Kusano
K
K
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Japan.
Abe
H
H
Obinata
T
T
D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002645:Chickens; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid
D051339:Actin Depolymerizing Factors; D000199:Actins; D008840:Microfilament Proteins; D010716:Phosphatidylinositols; D011994:Recombinant Proteins