通过基于结构的抗体工程提高治疗性抗体对甲基苯丙胺和苯丙胺的亲和力。
Affinity improvement of a therapeutic antibody to methamphetamine and amphetamine through structure-based antibody engineering.
作者信息
Thakkar Shraddha, Nanaware-Kharade Nisha, Celikel Reha, Peterson Eric C, Varughese Kottayil I
机构信息
Department of Physiology and Biophysics, College of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA.
Department of Pharmacology and Toxicology, College of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA.
出版信息
Sci Rep. 2014 Jan 14;4:3673. doi: 10.1038/srep03673.
Methamphetamine (METH) abuse is a worldwide threat, without any FDA approved medications. Anti-METH IgGs and single chain fragments (scFvs) have shown efficacy in preclinical studies. Here we report affinity enhancement of an anti-METH scFv for METH and its active metabolite amphetamine (AMP), through the introduction of point mutations, rationally designed to optimize the shape and hydrophobicity of the antibody binding pocket. The binding affinity was measured using saturation binding technique. The mutant scFv-S93T showed 3.1 fold enhancement in affinity for METH and 26 fold for AMP. The scFv-I37M and scFv-Y34M mutants showed enhancement of 94, and 8 fold for AMP, respectively. Structural analysis of scFv-S93T:METH revealed that the substitution of Ser residue by Thr caused the expulsion of a water molecule from the cavity, creating a more hydrophobic environment for the binding that dramatically increases the affinities for METH and AMP.
甲基苯丙胺（METH）滥用是一个全球性威胁，目前尚无任何获得美国食品药品监督管理局（FDA）批准的药物。抗METH免疫球蛋白G（IgGs）和单链抗体片段（scFvs）在临床前研究中已显示出疗效。在此，我们报告了通过引入点突变来增强一种抗METH scFv对METH及其活性代谢物苯丙胺（AMP）的亲和力，这些点突变是经过合理设计的，旨在优化抗体结合口袋的形状和疏水性。使用饱和结合技术测量结合亲和力。突变体scFv-S93T对METH的亲和力提高了3.1倍，对AMP的亲和力提高了26倍。scFv-I37M和scFv-Y34M突变体对AMP的亲和力分别提高了94倍和8倍。scFv-S93T与METH的结构分析表明，丝氨酸（Ser）残基被苏氨酸（Thr）取代导致一个水分子从腔中排出，为结合创造了一个更疏水的环境，从而显著增加了对METH和AMP的亲和力。
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