Mapping of drug binding sites on al-acid glycoprotein

人类酸性糖蛋白至少有两种遗传变异(A和F1*S变异)，它们由两个不同的基因编码。AGP是流通中基本药物的主要载体，不同的AGP变异体具有不同的药物结合特性。本研究的目的是利用定点突变技术鉴定与AGP基因变异的药物结合选择性有关的氨基酸残基。首先，我们用超滤技术筛选了与华法林、潘生丁和普罗帕酮结合的第100位氨基酸残基。华法林和潘生丁是F1*S的特异性配体，普罗帕酮是A特异性的配体。在F1*S变异体中，His97和His100分别参与了华法林和潘生丁的结合；Trp122也参与了这两种配体的结合。G1u92、His100和TrP122参与了普罗帕酮与A变异体的结合。AGP突变体之间第92位残基的交换逆转了普罗帕酮与两个突变体结合的相对强度，但对华法林和潘生丁的结合有明显不同的影响。V92E突变降低了华法林与F1*S突变体的结合，而E92V突变增加了双嘧达莫与A突变体的结合；尽管这两种药物与野生型F1*S突变体的结合亲和力都高于这两个突变体。这些发现表明，第92位氨基酸残基在AGP变异体的药物结合选择性中起着重要作用，特别是对优先与A变异体结合的药物。

There are at least two genetic variants of human al-acid glycoprotein (AGP) (the A and F1*S variants)that are encoded by two different genes. AGP is a major carrier of basic drugs in circulation and the variants of AGP have different drug-binding properties. The purpose of this study was to identify the amino acid residues that are responsible for the selectivity of drug binding to genetic variants of AGP using site-directed mutagenesis. First, we screened amino acid residues in the region proximal to position 100 that are involved in binding of warfarin and dipyridamole, which are F1*S-specific ligands, and of propafenone, which is an A-specific ligand, using ultrafiltration. In the F1*S variant, His97 and His100 were involved in warfarin- and dipyridamole-binding, respectively; Trp122 also contributed to binding of both ligands. G1u92, His100 and Trp 122 participated in the binding of propafenone in the A variant. Exchange of the residue at position 92 between AGP variants reversed the relative strength of propafenone binding to the two variants but had a markedly di8erent effect on binding of warfarin and dipyridamole. The V92E mutation decreased warfarin binding to the Fl*S variant, while the E92V mutation increased dipyridamole binding to the A variant; although, both drugs had greater binding affinities for the wild-type F1*S variant than for either mutant. These findings indicate that the amino acid residue at position 92 plays a significant role in drug-binding selectivity in AGP variants, especially for drugs that preferentially bind to the A variant.