The human alpha 2-macroglobulin receptor: identification of a 420-kD cell surface glycoprotein specific for the activated conformation of alpha 2-macroglobulin.

J D Ashcom,W S Argraves,J L Cravens,S E Tiller,D K Strickland,K Dickerson

doi:10.1083/jcb.110.4.1041

Abstract

Ligand affinity chromatography was used to purify a cell surface alpha 2-macroglobulin (alpha 2M) receptor. Detergent extracts of human placenta were applied to an affinity matrix consisting of alpha 2M, previously reacted with methylamine, coupled to Sepharose. Elution with EDTA specifically released polypeptides with apparent molecular masses of 420 and 39 kD. In some preparations, small amounts of a 90-kD polypeptide were observed. The 420- and 39-kD polypeptides appear specific for the forms of alpha 2M activated by reaction with proteinases or methylamine and do not bind to an affinity matrix consisting of native alpha 2M coupled to Sepharose. Separation of these two polypeptides was accomplished by anion exchange chromatography, and binding activity was exclusively associated with the 420-kD polypeptide. The purified 420-kD protein binds to the conformationally altered forms of alpha 2M that are known to specifically interact with alpha 2M receptors and does not bind to native alpha 2M. Binding of the 420-kD polypeptide to immobilized wheat germ agglutinin indicates that this polypeptide is a glycoprotein. The cell surface localization of the 420-kD glycoprotein was confirmed by affinity chromatography of extracts from surface radioiodinated fibroblasts. These properties suggest that the 420-kD polypeptide is a cell surface receptor for the activated forms of alpha 2M.

Highlights

Ligand affinity chromatography was used to purify a cell surface a2-macroglobulin receptor
Studies reported the specific binding of a2M to fibroblasts [9, 52], and it was recognized that the binding of ot2M to cells was specific for the forms of ~2M activated either by proteolysis or by reaction with methylamine [23, 32, 52]
Despite the widespread use of ot2Min studying the process of endocytosis, little is known about the polypeptide composition and structure of the o~2Mreceptor

Summary

Introduction

Ligand affinity chromatography was used to purify a cell surface a2-macroglobulin (ot2M) receptor. The 420- and 39-kD polypeptides appear specific for the forms of ~2M activated by reaction with proteinases or methylamine and do not bind to an affinity matrix consisting of native ot2M coupled to Sepharose. Separation of these two polypeptides was accomplished by anion exchange chromatography, and binding activity was exclusively associated with the 420-kD polypeptide. The reaction of proteinases or methylamine with c~2M"activates" the molecule by inducing conformational changes in o~2M(3, 4, 16, 49) that, in the case of the proteinase reaction, reduce the activity of the bound enzyme toward large molecular weight substrates These conformational changes generate sites on the inhibitor that result in specific recognition by cell surface receptors [31, 52, 54].

Methods

Results

Conclusion