Coactosin-like protein, a human F-actin-binding protein: critical role of lysine-75.

Coactosin-like protein (CLP) was recently identified in a yeast two-hybrid screen using 5-lipoxygenase as bait. In the present study, we report the functional characterization of CLP as a human filamentous actin (F-actin)-binding protein. CLP mRNA shows a wide tissue distribution and is predominantly expressed in placenta, lung, kidney and peripheral-blood ...
leucocytes. Endogenous CLP is localized in the cytosol of myeloid cells. Using a two-hybrid approach, actin was identified as a CLP-interacting protein. Binding experiments indicated that CLP associates with F-actin, but does not form a stable complex with globular actin. In transfected mammalian cells, CLP co-localized with actin stress fibres. CLP bound to actin filaments with a stoichiometry of 1:2 (CLP: actin subunits), but could be cross-linked to only one subunit of actin. Site-directed mutagenesis revealed the involvement of Lys(75) of CLP in actin binding, a residue highly conserved in related proteins and supposed to be exposed on the surface of the CLP protein. Our results identify CLP as a new human protein that binds F-actin in vitro and in vivo, and indicate that Lys(75) is essential for this interaction.
Mesh Terms:
Actins, Amino Acid Sequence, Animals, Binding Sites, Biopolymers, CHO Cells, COS Cells, Calcium, Carrier Proteins, Cricetinae, Cross-Linking Reagents, Female, Humans, Hydrogen-Ion Concentration, Lysine, Microfilament Proteins, Molecular Sequence Data, Neutrophils, Pregnancy, RNA, Messenger, Recombinant Proteins, Sequence Homology, Amino Acid, Tissue Distribution, Transfection, Two-Hybrid System Techniques
Biochem. J.
Date: Oct. 15, 2001
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