| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1994: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1993: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Research Abstract |
Bovine conglutinin is a Ca^<2+>-dependent serum lectin specific for N-acetylglucosamine and a member of the collectins, each of which possesses a collagenous region and carbohydrate binding domain (lectin) . In this study, we characterized the cDNA sequence and gene organization of conglutinin to shed light upon its evolutionary origin and and to elucidate its biological function with respect to the gene structure.
1) Firstly, the core region of conglutinin cDNA,which encoded the 175 amino acids from Pro28 to Gln203, was amplified by RT-PCR with degenerate oligonucleotide primers designed from the partial amino acid sequences we had determined previously and bovine liver poly (A) ^+ RNA as a template. Then, the 5'-and 3'-end regions of the conglutinin cDNA were amplified by the nested RACE.The amplified cDNA fragments of the core, 5'-, and 3'-end region was subcloned and the inserts were sequenced by the dideoxy method. The full-length cDNA (1548bp) structure was determined.
2) The conglutinin gene fragments were amplified by PCR with specific primers designed from the cDNA sequence using bovine liver genomic DNA as a template. The amplified DNA fragments were directly sequenced. The conglutinin gene spanned over 7.5kb, and the coding region of conglutinin mRNA consisted of seven exous. The long collagen-like domain (55 Gly-X-Y repeats) , which is characteristic of conglutinin, was encoded by five separate exons. The neck region and the carbohydrate-recognition domain were encoded by separate exons, respectively. The overall exon-intron organization of conglutinin was very similar to those of other collectins such as lung surfactant apoprotein D and mannan (mannose) -binding proteins, suggesting that these lectins are evolutionarily close and formed by a shuffling of exons derived from the collagen and carbohydrate-binding protein families.
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