Isolation of a Zn-binding protein mediating cell adhesion from common carp.
Extraordinarily high concentrations of Zn (300-500 microg/[g fresh tissue]) are often found in the digestive tract tissue of common carp Cyprinus carpio, and most of the Zn is bound to membrane protein located on plasma membranes that are attached to basal laminae. To isolate the Zn-binding protein, the basolateral plasma membranes were separated from the extracellular matrix by treating the nuclei/cell debris fraction of the tissue with collagenase type IV and Arg-Gly-Asp (RGD) peptide. The Zn-binding protein was isolated from the separated plasma membranes by immobilized metal affinity chromatography and affinity chromatography on laminin-Sepharose. A 43 kDa protein was bound by the laminin-Sepharose and specifically eluted with tirofiban (a mimic of RGD). Affinity chromatography on wheat germ agglutinin and concanavalin A-Sepharose showed that the 43 kDa protein is a glycoprotein. The 43 kDa protein was labelled with 65Zn and became incorporated into liposomes at a high efficiency. Liposomes containing this protein were bound to laminin-Sepharose or reconstituted basement membrane. We propose that the Zn-binding protein is a cell surface receptor involved in the adhesion of cells to laminin.
D000818:Animals; D001345:Autoradiography; D002347:Carps; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000502:physiology; D002448:Cell Adhesion / Q000502:physiology*; D002846:Chromatography, Affinity; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D008081:Liposomes; D009842:Oligopeptides / Q000494:pharmacology; D000077466:Tirofiban; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000494:pharmacology
D002352:Carrier Proteins; D003902:Detergents; D008081:Liposomes; D009842:Oligopeptides; C017800:zinc-binding protein; D014443:Tyrosine; C047981:arginyl-glycyl-aspartic acid; D000077466:Tirofiban