Physical characterization of hFSH and its desialylation products by isoelectric focusing and electrophoresis in polyacrylamide gel.

Abstract

Iodinated and tritiated derivatives of highly purified hFSH were characterized by polyacrylamide gel electrophoresis (PAGE) at pH 7.8 and 10.2, 0 C, and by isoelectric focusing in polyacrylamide gel (IFPA). 125I— and 3HhFSH appear homogeneous in PAGE, as evidenced by single, linear Ferguson plots at the two pH values, and by a single peak in IFPA. 131I—hFSH in contrast appears heterogeneous. The physical characteristics of hFSH, recognized most comprehensively by the parameters KR (retardation coefficient, a measure of molecular size) and Yo (a measure of molecular net charge) in PAGE, differed significantly between 3H— and 125I—hFSH: it appeared that the tritiated hormone was in an aggregated form at pH 10.2. However, the molecular size of 3H—hFSH appeared the same at pH 7.8 as that of 125I—hFSH at pH 10.2: both values correspond to a mol wt of 36,000, in agreement with literature values. Size discrepancy between 2 variously labeled preparations under a single set of PAGE conditions implies that recognit...