PURIFICATION, CLONING, AND SEQUENCE-ANALYSIS OF A M(R)=30,000 PROTEINFROM SEA-URCHIN AXONEMES THAT IS IMPORTANT FOR SPERM MOTILITY - RELATIONSHIP OF THE PROTEIN TO A DYNEIN LIGHT-CHAIN

We have generated a series of monoclonal antibodies against axonemal p roteins from sea urchin spermatozoa in order to identify novel protein s involved in the regulation of flagellar motility. The monoclonal ant ibody D405-14 inhibited the motility of demembranated-reactivated sper m models at low concentrations and recognized a single polypeptide of 33 kDa (p33) on immuno blots of sea urchin axonemal proteins. Fraction ation of the axonemes with high salt solutions, heat, and detergent re sulted in the selective extraction of p33 into a 0.6 M NaCl-soluble an d a 0.5% sodium lauryl sarcosinate (Sarkosyl)-soluble form. Both forms of p33 were purified to apparent homogeneity by immunoaffinity chroma tography on monoclonal antibody D405-14-Sepharose. We have also isolat ed and sequenced a full-length cDNA clone encoding the 33-kDa protein. The sequence predicts a polypeptide of 260 amino acids having a mass of 29,730 Da and an isoelectric point of 9.3. Sequence comparison indi cates that p33 is 66% identical (74% similar) to the p28 light chain o f axonemal inner dynein arm of Chlamydomonas reinhardtii. Taken togeth er, these results suggest that we have identified a p28 light chain ho molog in sea urchin sperm axoneme and that this protein may play a dyn amic rob in flagellar motility.