PURIFICATION AND CHARACTERIZATION OF 26S PROTEASOME FROM SPERM FLAGELLA OF CHUM SALMON AND ITS ROLES IN THE REGULATION OF SPERM MOTILITY

A protease with molecular mass of 1,400 kDa has been purified from chu m salmon sperm flagella in the presence of ATP. The purified enzyme ex hibited multicatalytic protease activity towards peptide substrates fo r trypsin-like, chymotrypsin-like and peptidylglutamylpeptide hydrase- like proteases. The enzyme was composed of at least 38 subunits in the range of 21-120 kDa on SDS-PAGE, the pattern of which was quite simil ar to that of 26S proteasome found in many eukaryotic cells. The subun its in the range of 21-32 kDa were identical to those of the 20S prote asome, which was previously purified from chum salmon sperm flagella ( 12). These results suggest that the 1,400 kDa protease is the 26S form of proteasome in chum salmon sperm flagella. The 26S proteasome subun its in the range of 35-120 kDa were revealed to be mostly identical to those of the 950 kDa protease (15), suggesting that the 26S proteasom e is formed by ATP-dependent association of the 20S proteasome and the 950 kDa protease. The peptide substrates of the 26S proteasome and pr oteasome inhibitors inhibited the motility of demembranated sperm of c hum salmon. The inhibition became remarkable with an increase in ATP c oncentration. These suggest that flagellar 26S proteasome plays a key role in the regulation of flagellar motility of spermatozoa in salmoni d fish.