|Budget Amount *help
¥6,400,000 (Direct Cost : ¥6,400,000)
Fiscal Year 1991 : ¥1,300,000 (Direct Cost : ¥1,300,000)
Fiscal Year 1990 : ¥5,100,000 (Direct Cost : ¥5,100,000)
Human serum mannan-binding protein (MBP) is a lectin (carbohydrate binding protein), which was first described by these authors. The lectin has been shown to have the ability to activate complement and also to function as direct opsonin, leading to the concept that the lectin is a novel component associated with selfdefense mechanism. This study was undertaken to examine the structure function relationship particularly on its complement activating activity of the lectin with the results described below.
1) Amino acid sequence of the human serum MBP was studied protein chemically. The lectin, which had been reduced and alkylated, was digested with lysylendopeptidase. The resulting peptides were purified by HPLC and then their sequences were determined by a peptide sequencer.
2) Human serum MBP cDNA was cloned from human liver cDNA library, which had been carried in lambda gtll, by screening with rat serum MBP cDNA as a probe and the nucleotide sequence of the cloned cDNA was determined by
dedeoxy termination method. By comparing the result obtained in 1) and 2), human serum MBP was revealed to be composed of 248 amino acids.
3) The gross structure of the serum MBP is very similar to a complement component Clq. This prompted to us to study the molecular mechanism of complement activation by the serum MBP. Studies using the other complement components Clr and Cls revealed that the serum MBP, once bound to the solid phase ligands, associates Clr*Cls complement. On this complex, the conversion of Clr*Cls from their zymogen forms to the activated serine proteases proceeds, leading to the activating of a series of cascade reaction of the classical pathway of the complement system.
4) The serum MBP was shown to have complement dependent bactericidal activity against E. coli K12, B strains which have haptenic sugars (mannose, NAcetylglucosamine) on their core structure, and also cytotoxic activity against cultured mammalian cells which have high mannose-type oligosaccharide chains on their surfaces.
5)血清MBPの補体活性化に関与する部位を同定するため,MBPの各部分構造に対応する合成ペプチドを調製し,MBPによる補体活性化反応の阻害活性を調べたところ,コラ-ゲン様ドメインのC未端側lC Clr・Cls結合部位が存在することが明らかとなった。 Less