Glycoproteins present in human follicular fluid that inhibit the zona- binding capacity of spermatozoa

Previous studies have suggested that human follicular fluid containsfactors that reduce the zona-binding capacity of spermatozoa. The presentstudy provides further evidence of the existence of such factors. Using thehemizona binding assay (HZA), we have shown that the inhibitory effect ofhuman follicular fluid on the zona-binding capacity of spermatozoa isconcentration-dependent, an inhibitory effect being detected when theconcentration of human follicular fluid was > or = 10%. A 1%concentration of human follicular fluid did not possess this inhibitoryactivity. Heating human follicular fluid at 56 degrees C for 30 min did notaffect its inhibitory properties; treatment with proteinase-K abolishedsuch inhibition. Human follicular fluid was fractionated sequentially byconcanavalin-A affinity chromatography, Mono Q ion-exchange chromatographyand Superose-12 gel filtration. The zona binding inhibitory activityresided in the fraction which bound to the lectin and Mono Q column andcontained molecules with native molecular weights of 32 and 192 kDa. Sodiumdodecyl sulphate-polyacrylamide gel electrophoresis analysis suggested thatthe 192 kDa glycoprotein was a tetramer, while the 32 kDa glycoproteinremained as a single molecular species under denaturing conditions. Weconclude that two glycoproteins were responsible for the zona bindinginhibitory activity of human follicular fluid. The physiological role ofthese factors remains unclear.