Molecular cloning and sequence analysis of aggretin, a collagen-like platelet aggregation inducer.
A cDNA library derived from the Malayan-pit-viper (Calloselasma rhodostoma) venom gland was constructed in the phagemid vector. Using the information of the N-terminal amino acid sequences of two subunits of aggretin, synthetic mixed-base oligonucleotides were employed as a screening probe for colony hybridization. Separate cDNA clones encoding for the alpha and beta chains of aggretin were isolated and sequenced. The results revealed that mature alpha and beta chains contain 136 and 123 amino acid residues, respectively. Aggretin subunits show high degrees of identity with respective subunits (50-60% for alpha, 49-58% for beta) of C-type lectin-like snake venoms. The identity to rattlesnake lectin is relatively lower (i.e., 39 and 30%). All cysteine residues in each chain of aggretin are well conserved and located at the positions corresponding to those of C-type lectins. Thus, three intracatenary disulfide bridges and an interchain disulfide bond between Cys83(alpha) and Cys75(beta) may be allocated. This is the first report regarding the entire sequence of venom GPIa/IIa agonist. According to the alignment of amino acid sequences, hypervariable regions among these C-type lectin-like proteins were revealed. These hypervariable regions are proposed to be the counterparts directly interacting with different receptors or different domains of a receptor on the surface of platelet.
Chung
C H
CH
College of Medicine, National Taiwan University, Taipei, Taiwan.
Au
L C
LC
Huang
T F
TF
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003094:Collagen / Q000494:pharmacology; D017839:Crotalus / Q000235:genetics; D015723:Gene Library; D037102:Lectins / Q000737:chemistry; D037181:Lectins, C-Type; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D010974:Platelet Aggregation / Q000187:drug effects*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012910:Snake Venoms / Q000737:chemistry; D014757:Viper Venoms / Q000737:chemistry* / Q000235:genetics* / Q000494:pharmacology
D037102:Lectins; D037181:Lectins, C-Type; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D012910:Snake Venoms; D014757:Viper Venoms; C112292:rhodocytin protein, Calloselasma rhodostoma; D003094:Collagen