Purification and characterization of zyxin, an 82,000-dalton component of adherens junctions.
We describe here the purification and characterization of a recently identified adherens junction protein that has an apparent molecular mass of 82 kDa on sodium dodecyl sulfate-polyacrylamide gels (Beckerle, M. C. (1986) J. Cell Biol. 103, 1679-1687). The 82-kDa protein was isolated from avian smooth muscle by a low ionic strength alkaline pH extraction followed by ammonium sulfate fractionation. Sequential chromatographic separation using DEAE-cellulose, phenyl-Sepharose CL-4B, and hydroxylapatite resins results in a purified 82-kDa protein. The 82-kDa protein has a Stokes radius of 5.6 nm and a relative sedimentation coefficient of 3.0 S. The calculated native molecular mass of the protein based on its hydrodynamic properties is 69 kDa, and the derived frictional ratio (f/fo) is 2.1. The protein does not focus discretely by isoelectric-focusing-sodium dodecyl sulfate-polyacrylamide gel electrophoresis; there are numerous isoelectric point variants in the range of 6.4-7.2, with the average isoelectric point being 6.9. The 82-kDa protein is phosphorylated in vivo and appears to be a cytoplasmic component of adherens junctions. The properties of the 82-kDa protein distinguish it from other known adherens junction proteins of this molecular mass. In fibroblasts, the 82-kDa protein is found in adhesion plaques as well as along actin-containing stress fibers near where they terminate at sites of cell-substratum adhesion. It is also found in the cell-cell adherens junctions of pigmented retinal epithelial cells and the dense plaques of smooth muscle cells. Since the 82-kDa protein is found at both cell-substratum and cell-cell adherens junctions, we propose to call it zyxin, meaning a joining, to indicate that it is found at regions where extracellular ligands are structurally and functionally joined to the cytoskeleton.
Crawford
A W
AW
Department of Biology, University of Utah, Salt Lake City 84112.
Beckerle
M C
MC
D000818:Animals; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D007365:Intercellular Junctions / Q000378:metabolism*; D007526:Isoelectric Point; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D009130:Muscle, Smooth / Q000378:metabolism* / Q000648:ultrastructure; D010766:Phosphorylation
D015815:Cell Adhesion Molecules; D008565:Membrane Proteins