Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P04626 (erbB-2)
 5,251 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have used a variety of methods, including lactoperoxidase-catalyzed iodination, proteolysis, and photolabel incorporation, to determine whether exposure to the acidic pH encountered during receptor-mediated endocytosis causes observable conformational changes in receptor proteins. Two receptor systems were chosen for this study: the asialoglycoprotein receptor and the epidermal growth factor (EGF) receptor. The purified asialoglycoprotein receptor protein was reconstituted into lipid membranes by spontaneous incorporation into phosphatidylcholine liposomes with the binding site facing outward. The EGF receptor was studied in living A-431 cells and was identified by immunoprecipitation using monoclonal antibodies. Lactoperoxidase-catalyzed iodination of both receptor systems, carried out with the external pH equal to 7.4 or 5.6, showed that the extent of receptor protein iodination was less at the lower pH even though lactoperoxidase has an acidic pH optimum. Using the nonspecific hydrophilic photolabeling agent [35S]N-(4-azido-3-nitrophenyl)-2-aminoethylsulfonic acid-taurine, we observed less incorporation into both the asialoglycoprotein receptor in liposomes and the EGF-receptor in A-431 cells when the external pH was reduced to 5.6. Also, using the enzyme papain, we have found that both receptors become resistant to proteolysis when the external pH is lowered from 7.0 to 5.6. These results suggest a conformational change in both of these receptors in which they become less exposed to the external aqueous environment at low pH. Such a conformational change may be responsible for the pH dependence of binding for both of these ligands. Also, this conformational change may serve to protect receptors from enzymatic degradation within endocytic or lysosomal compartments.
 ...
PMID:Conformational changes in the receptors for epidermal growth factor and asialoglycoproteins induced by the mildly acidic pH found in endocytic vesicles. 608 23

The membranes from epidermoid carcinoma cells (A-431) that were surface iodinated while intact using catalysis by lactoperoxidase and 125I as iodide contain one major labeled protein of Mr = 180,000. This protein is clearly iodinated on the outside of the intact cell because it is not the major protein labeled when isolated membranes are iodinated. This major surface-iodinated protein is almost certainly the epidermal growth factor (EGF) receptor, since both have the same Mr and have identical sensitivity to proteases. Both are nearly quantitatively converted from an Mr = 180,000 form to an Mr = 160,000 form by an endogenous calcium-activated neutral protease when cells are broken in the presence of calcium. Both are degraded by trypsin only if trypsin has access to the inside of the cell. This latter finding implies that the surface-iodinated EGF receptor spans the plasma membrane. Since the EGF receptor is an autophosphorylating kinase whose activity is enhanced in the presence of EGF, the receptor was labeled and identified using [gamma-32P] ATP. While both iodination and EGF-enhanced phosphorylation occur on tyrosine residues, peptide mapping of the iodinated or phosphorylated Mr = 180,000 band showed that different peptides were being labeled. Since the EGF receptor-kinase spans the plasma membrane, the peptide iodinated on the surface of the intact cell must be different from the peptides that are probably autophosphorylated on the cytoplasmic side of the membrane.
 ...
PMID:Surface iodination of epidermal growth factor receptors in intact cells. 632 89