Little R G, Kelner D N, Lim E, Burke D J, Conlon P J
Department of Immunology, XOMA Corporation, Berkeley, California 94710.
J Biol Chem. 1994 Jan 21;269(3):1865-72.
The 23-kDa recombinant amino-terminal bactericidal/permeability increasing protein fragment (rBPI23) has all of the antibacterial and antiendotoxin properties of the holoprotein. In the current studies, we have identified multiple active domains within rBPI23 with chemical and proteolytic cleavage fragments and with synthetic overlapping peptides. We also demonstrate a novel, high affinity heparin binding property for rBPI23, in addition to its established bactericidal and lipopolysaccharide binding properties. Cleavage fragments and synthetic, overlapping peptides of rBPI23 were analyzed for inhibition of the lipopolysaccharide-induced Limulus amebocyte lysate reaction, for bactericidal activity, and for heparin binding. Three separate, active domains were identified in amino acid regions 17-45, 65-99, and 142-169. A single synthetic peptide (85-99) was bactericidal. These results indicate that rBPI23 is comprised of three separate functional domains which contribute to the high affinity interaction of rBPI23 with Gram-negative bacteria. The individual activity of each domain and the cooperative interaction among domains provide the basis for developing rBPI23 analogues with increased biologic efficacy.
23 kDa重组氨基末端杀菌/通透性增加蛋白片段（rBPI23）具有完整蛋白的所有抗菌和抗内毒素特性。在当前研究中，我们通过化学和蛋白水解裂解片段以及合成重叠肽确定了rBPI23内的多个活性结构域。除了已确定的杀菌和脂多糖结合特性外，我们还证明了rBPI23具有一种新的高亲和力肝素结合特性。对rBPI23的裂解片段和合成重叠肽进行了分析，以检测其对脂多糖诱导的鲎试剂反应的抑制作用、杀菌活性和肝素结合能力。在氨基酸区域17 - 45、65 - 99和142 - 169中确定了三个独立的活性结构域。单个合成肽（85 - 99）具有杀菌作用。这些结果表明，rBPI23由三个独立的功能结构域组成，这些结构域有助于rBPI23与革兰氏阴性菌的高亲和力相互作用。每个结构域的个体活性以及结构域之间的协同相互作用为开发具有更高生物学效能的rBPI23类似物提供了基础。
