Inhibition of flagellar motility of demembranated fowl spermatozoa by protease substrates.

We investigated the effects of various protease substrates on the motility of demembranated fowl spermatozoa. In the presence of ATP, the motility of demembranated spermatozoa was vigorous at 30 degrees C, but decreased markedly following the addition of protease substrates, such as N alpha-carbobenzoxy-L-lys-thiobenzyl ester (BLT), N-benzoyl-phe-val-arg p-nitroanilide or N alpha-benzoyl-D,L-arg p-nitroanilide (BAPNA) in a dose-dependent manner, within the range 0-1 mM. The subsequent addition of 100 ng/ml trypsin released the inhibitory effect of protease substrates within 10 s. Phosphorylation or dephosphorylation of several proteins of demembranated spermatozoa was observed following the addition of protease substrates, however, no consistent patterns of protein phosphorylation or dephosphorylation were associated with the inhibition of motility. These results suggest that endogenous protease activity is instrumental in the maintenance of fowl sperm motility and that the site of action of this protease is in the axoneme and/or accessory cytoskeletal components. This enzyme may not act directly on the phosphorylation of sperm proteins involved in the regulation of motility.