Theoretical Model for HIV‑1 PR That Accounts for Substrate Recognition and Preferential Cleavage of Natural Substrates

The
Human Immunodeficiency Virus type 1 (HIV-1) protease is a crucial
target for HIV/AIDS treatment, and understanding its catalytic mechanism
is the basis on which HIV-1 enzyme inhibitors are developed. Several
experimental studies have indicated that HIV-1 protease facilitates
the cleavage of the Gag and Gag-Pol polyproteins and it is highly
selective with regard to the cleaved amino acid precursors and physical
parameters. However, the main theoretical principles of substrate
specificity and recognition remain poorly understood theoretically.
By means of a one-step concerted transition state modeling, the recognition
of natural substrates by HIV-1 PR subtypes (B and C-SA) was studied.
This was carried out to compare the activation free energies at varying
peptide bond regions (scissile and nonscissile) within the polypeptide
sequence using ONIOM calculations. We studied both P3–P3′
and P5–P5′ natural substrate systems. For P3–P3′
substrates, excellent recognition was observed for the MA-CA family
but not for the RH-IN substrates. Satisfactory recognition for the
latter was only observed for the longer sequence (P5–P5′)
after the substrate was subjected to an MD run to maximize the interaction
between the enzyme and the substrate. These results indicate that
both sequence and structure are important for correct scissile bond
recognition of these natural substrates.

人类免疫缺陷病毒1型（HIV-1）蛋白酶是艾滋病（HIV/AIDS）治疗的关键靶点，阐明其催化机制是开发HIV-1蛋白酶抑制剂的核心基础。多项实验研究表明，HIV-1蛋白酶可催化Gag与Gag-Pol多聚蛋白的水解切割，且对切割位点的氨基酸前体及物理参数具有高度选择性。然而，底物特异性与识别的核心理论原理在理论层面仍未得到充分阐释。本研究采用一步协同过渡态建模方法，针对HIV-1蛋白酶（PR）的B亚型与C-SA亚型的天然底物识别行为展开研究：通过ONIOM计算，对比多肽序列中不同肽键区域（可切割位点与非可切割位点）的活化自由能。我们同时考察了P3-P3′与P5-P5′两类天然底物体系。针对P3-P3′底物，MA-CA家族底物展现出优异的识别效果，但RH-IN底物则未观察到该现象。仅当对底物进行分子动力学（MD）运行以最大化酶与底物间的相互作用后，更长序列的P5-P5′型RH-IN底物才获得了令人满意的识别效果。上述结果表明，序列特征与空间结构对于这类天然底物的可切割键正确识别均具有重要意义。