STRUCTURE OF RECEPTORS FOR ENDOCYTOSIS OF GLYCOPROTEINS

The experiments proposed are designed to contribute to a broad-based understanding of the molecular basis for receptor-mediated endocytosis in eukaryotic cells and the role of complex carbohydrates in biological systems. Work will be concentrated on a specific group of hepatocyte plasma membrane receptors which recognize partially deglycosylated serum glycoproteins; these proteins are selected for study because they can be purified in sufficient quantities for structural studies. The receptors are referred to as hepatic lectins. The work proposed covers the following areas: (a) amino acid sequence analysis of the minor species of the rat hepatic lectin as well as comparison of the functional properties of the several species of this receptor to determine the specific role of each of these homologous but distinct proteins; (b) structure-function studies on the chicken and rat receptors to identify residues which form the ligand binding sites, and to define as precisely the portions of these proteins which interact with the lipid bilayer; (c) detailed analysis of the biosynthesis of the receptor polypeptides, with emphasis on determining the mechanism of insertion into a membrane, using the techniques of in vitro translation and molecular cloning; and (d) comparative study, including primary structure analysis, of the mannose- specific receptor from rat liver to determine if this protein is related to the membrane lectins, studied to this point or, as seems more likely from recent results, if this receptor shares structural and function properties with complement protein Clq. As a part of the primary structure studies on membrane proteins, investigation of the sequence of a variant form of the erythrocyte membrane anion transport protein is also proposed. These studies would exploit the availability of pure receptors to advance our understanding of the mechanism of endocytosis; in addition, the hepatic lectins are models for receptors involved in other processes such as hormone signalling and cell-cell recognition. The analysis of the several proteins described will also help to elucidate the function of complex carbohydrate in biological recognition processes and will provide detailed information about the mechanism of membrane biogenesis.