Study of the interaction of cumulus-associated alpha-2-macroglobulin with glycodelin-C from human cumulus matrix

Abstract

Glycodelin is a glycoprotein having 4 isoforms: glycodelin-A (GdA, amniotic fluid isoform), glycodelin-S (GdS, seminal plasma), glycodelin-F (GdF, follicular fluid) and glycodelin-C (GdC, cumulus matrix) with molecular sizes ranging from 25-30 kDa. Glycodelin isoforms have identical protein core but different glycosylation, which determines their biological activities. In contrast to the spermatozoa-zona pellucida (ZP) binding inhibitory activity of GdA and GdF, GdC from the cumulus cell matrix enhances spermatozoa-ZP binding. In this study, we purified a glycodelin-C binding protein (F3) from the human cumulus matrix with molecular size of 30 kDa using anti-glycodelin immunoaffinity chromatography followed by ion-exchange chromatography. The protein was identified to be a member of alpha-2-macroglobulin (A2MG) protein family by MS/MS-TOF analysis. The granulosa cells, but not the cumulus cells, expressed A2MG mRNA. Moreover, our data suggested that the cumulus cells could convert A2MG into a smaller molecule with size similar to that of F3. The interaction between F3 and GdC was carbohydrate dependent as evidence by ability of F3 to bind to GdC but not to deglycosylated glycodelin. F3 also interacted with hyaluronic acid, a main component of the cumulus matrix. In contrast to GdC, F3 did not bind to human spermatozoa. We proposed that the cumulus cells converted granulosa cell-derived A2MG into F3 and deposited into the cumulus matrix. GdC was retained in the matrix by binding to F3 which was held in position by binding to hyaluronic acid. Further works are required to characterize the role of F3 in the biological activities of GdC on human spermatozoa. This work was supported in part by grants from the Hong Kong Research Grant Council (HKU7261/01M, HKU7408/03M and HKU7647/06M),