Abstract:
Crotaline venoms contain a variety of C-type lectin-like proteins that participate in hematotoxicity. Previously characterized lectins show conserved folds with several disulfide bridges. The structures are composed of alfa beta hetero-oligomers linked by interchain disulfide bonds. They may enhance or inhibit platelet-ligand bindings, as well as inhibition of clotting factor IX and X. In previous studies, the platelet glycoproteinIb-binding protein from trimeresurus albolabris named alboaggregin B (AL-B), has been purified and sequenced. However, there is no report of the cDNA sequences of AL-B. The aim of this thesis is to analyze the full-length sequence of AL-B alfa and beta subunits from the cDNA library of T. albolabris venom gland. Moreover the protein has been purified and characterized. Based on partial sequence of the primary library, oligonucleotide primers were designed and used to clone full-length cDNA encoding the C-type lectin subunit by 5'-RACE method. Additional homologous subunit was obtained using 3'-RACE. Analysis of the full-length nucleotide sequence found that the coding region of AL-B alfa subunit was 471 bp and beta subunit was 441 bp with the deduced protein sequence of 133 and 123 amino acid residues respectively, including 23 residues signal peptides. The nucleotide sequecne of AL-B alfa and beta subunit showed 85.25% and 79.45% amino acid sequence identity with stejaggregin from T. stejnegeri, respectively. In addition, by sequence comparison a conserved motif in AL -B beta (SRTY) that may be responsible for platelet aggregation activity was found. Furthermore, AL-B protein was purified from the snake venom and confirmed to be identical to the cloned protein using the MALDI TOF mass spectroscopy. AL-B agglutinated fixed human platelets with the EC50 of 180 nM and was completely inhibited by anti GPIb antibody. To our knowledge, this is the first report of cDNA cloning of C-typr lectin-like protein from T. albolabris.
