Preventing the spread of infectious diseases remains an urgent priority worldwide, and this is driving the development of advanced nanotechnology to diagnose infections at the point of care. Herein, we report the creation of a library of novel nanobody capture ligands to detect p24, one of the earliest markers of HIV infection. We demonstrate that these nanobodies, one tenth the size of conventional antibodies, exhibit high sensitivity and broad specificity to global HIV-1 subtypes. Biophysical characterization indicates strong 690 pM binding constants and fast kinetic on-rates, 1 to 2 orders of magnitude better than monoclonal antibody comparators. A crystal structure of the lead nanobody and p24 was obtained and used alongside molecular dynamics simulations to elucidate the molecular basis of these enhanced performance characteristics. They indicate that binding occurs at C-terminal helices 10 and 11 of p24, a negatively charged region of p24 complemented by the positive surface of the nanobody binding interface involving CDR1, CDR2, and CDR3 loops. Our findings have broad implications on the design of novel antibodies and a wide range of advanced biomedical applications.

中文翻译：

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揭示针对 HIV p24 的高亲和力纳米抗体的分子基础：体外功能、结构和计算机洞察力

预防传染病的传播仍然是世界范围内的当务之急，这正在推动先进纳米技术的发展，以在护理点诊断感染。在这里，我们报告创建了一个新型纳米抗体捕获配体库来检测 p24，这是 HIV 感染的最早标志物之一。我们证明，这些纳米抗体的大小是常规抗体的十分之一，对全球 HIV-1 亚型表现出高灵敏度和广泛特异性。生物物理特性表明 690 pM 的强结合常数和快速的动力学结合率，比单克隆抗体比较器好 1 到 2 个数量级。获得了铅纳米体和 p24 的晶体结构，并与分子动力学模拟一起使用，以阐明这些增强性能特征的分子基础。他们表明结合发生在 p24 的 C 末端螺旋 10 和 11，p24 的带负电荷的区域由涉及 CDR1、CDR2 和 CDR3 环的纳米体结合界面的正表面补充。我们的研究结果对新型抗体的设计和广泛的先进生物医学应用具有广泛的影响。