Putative rat sperm lipid-binding protein: isolation and partial characterization.
Previous work has identified a prominent 22-24-kD protein that is present in rat male reproductive tissues, including epididymis and testis (Brooks, 1985; Jones and Brown, 1987; Moore et al., 1987). Using a monoclonal antibody (designated mAb-B109) against this 24-kD antigen (referred to as B109), we have isolated the protein using a combination of chromatofocusing and electroelution from SDS-PAGE gels, and reverse phase HPLC. B109 (pI = 4.8) is amino-terminal blocked. To obtain internal amino acid sequences, the isolated protein was cleaved either with cyanogen bromide in 70% formic acid or with TLCK-treated chymotrypsin. With cyanogen bromide treatment, two peptides, 17.8 kD and 11.9 kD, were isolated and partial amino acid sequences obtained. Chymotryptic peptides were isolated by reverse-phase HPLC and two were chosen for sequence analysis. A computer search for sequence homology through the protein identification resource (PIR) matched B109 to a basic 21-kD cytosolic protein (pI = 7.4) found in bovine brain (> 80% homology). When peptide sequence differences obtained in the present study were substituted into the 21-kD cytosolic protein sequence obtained from the PIR using Intelligenetics software, the calculated pI dropped from 7.4 to 5.8, suggesting that pI differences between the bovine and rat molecules are the result of amino acid substitutions in the testis protein and not tissue-specific posttranslational processing. It has been postulated that the 21-kD bovine brain protein is associated with phospholipid transport, although the function of B109 is unknown.
Pryor
J L
JL
Department of Cell Biology and Neuroanatomy, University of Minnesota, Minneapolis.
Xu
W
W
Moore
A
A
Ensrud
K M
KM
Hamilton
D W
DW
D000595:Amino Acid Sequence; D000818:Animals; D000941:Antigens / Q000302:isolation & purification* / Q000378:metabolism; D001923:Brain Chemistry; D002352:Carrier Proteins; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D050356:Lipid Metabolism; D008297:Male; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017386:Sequence Homology, Amino Acid; D013094:Spermatozoa / Q000737:chemistry*
D000941:Antigens; C092593:B109 protein, rat; D002352:Carrier Proteins; D011506:Proteins