Biochemical Characterization of Two Different Forms of the Substance P Receptor in Rat Submaxillary Gland

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Abstract
Studies were designed to examine the basis for the difference in molecular weights of the two proteins detected in membrane preparations of rat submaxillary glands after photolabeling with a radioactive analogue of substance P, 125I-p-benzoyl-L-phenylalanine8-substance P. When the two proteins were separated and individually digested with endoglycosidase F, the relative molecular weight of each protein was reduced by ∼ 10,000, indicating that the extent of glycosylation of both proteins is the same. To test whether the difference in their molecular weights can be attributed to a difference in the lengths of the two proteins, photolabeled membranes were treated with carboxypeptidase Y before solubilization to remove from each photolabeled protein the carboxy-terminal portion that extends beyond the membrane. Only one, albeit diffuse, band was now observed that on subsequent deglycosylation with endoglycosidase F was more clearly seen to be a single band, indicating that differing lengths of peptide chains were cleaved from the two proteins. These results permit the interpretation that the difference in the two forms of the substance P receptor present in rat submaxillary glands is due to differences in the length of their carboxy termini.