ANALYSIS OF THE SHORT CONSENSUS REPEATS OF HUMAN-COMPLEMENT FACTOR-B BY SITE-DIRECTED MUTAGENESIS
ANALYSIS OF THE SHORT CONSENSUS REPEATS OF HUMAN-COMPLEMENT FACTOR-B BY SITE-DIRECTED MUTAGENESIS
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DOI：
10.1074/jbc.270.34.19716
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发表时间：
1995-08-25
影响因子：
4.8
通讯作者：
OGLESBY, TJ
中科院分区：
文献类型：
--
作者：
HOURCADE, DE;WAGNER, LM;OGLESBY, TJ
人因子B是补体替代途径的起始和繁殖所必需的，它也参与补体经典途径的扩增。单独来看，因子B是一种生化活性知之甚少的酶原，但在替代途径转化酶的背景下，因子B丝氨酸蛋白酶被激活的过程首先涉及与C3b结合，随后将因子B切割成两个片段，Ba和Bb， Ba，nh2末端片段，主要由三个串联的短一致重复组成，在其他补体蛋白中也发现球状结构域，它在组装过程中与转换酶分离，留下活跃的C3转换酶C3bBb，以前的报道表明Ba区域可能在转换酶组装中起作用。利用重组因子B的定点诱变和单克隆抗体表位定位来评估特异性短共识重复氨基酸残基的相对重要性，对这一假设进行了检验，确定了三个感兴趣的位点，位点1是短共识重复1中19个连续氨基酸的延伸，形成了有效阻断因子B功能的单克隆抗体的表位。位点2由短共识重复2中的6个连续氨基酸组成，位点3由短共识重复3中的7个连续氨基酸组成，突变使因子B溶血活性降低至3%或更低。进一步的分析表明，位点2和位点3与因子B- c3b相互作用有关。
Human factor B is required for the initiation and propagation of the complement alternative pathway, It also participates in the amplification of the complement classical pathway, Alone, factor B is a zymogen with Little known biochemical activity, but in the context of the alternative pathway convertases, the factor B serine protease is activated in a process that first involves the association with C3b and subsequently the cleavage of factor B into two fragments, Ba and Bb, Ba, the NH2-terminal fragment, is composed mainly of three tandem short consensus repeats, globular domains found in other complement proteins, It dissociates from the convertase during assembly, leaving the active C3 convertase, C3bBb, Previous reports suggest that the Ba region may be instrumental in convertase assembly, This hypothesis was tested using site-directed mutagenesis of recombinant factor B and monoclonal antibody epitope mapping to evaluate the relative importance of specific short consensus repeat amino acid residues, Three sites of interest were identified, Site 1 is a stretch of 19 contiguous amino acids in short consensus repeat 1 that form the epitope of a monoclonal antibody that effectively blocks factor B function. Site 2, composed of 6 contiguous amino acids in short consensus repeat 2, and site 3, consisting of 7 contiguous amino acids in short consensus repeat 3, were defined by mutations that reduce factor B hemolytic activity to 3% or less, Further analyses indicated that sites 2 and 3 contribute to factor B-C3b interactions.