计算设计的小蛋白受热引起的结构和化学变化。

IF 4.5 3区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY
Protein Science Pub Date : 2024-06-01 DOI:10.1002/pro.4991
Joshua A Dudley, Sojeong Park, Oliver Cho, Nicholas G M Wells, Meagan E MacDonald, Katerina M Blejec, Emmanuel Fetene, Eric Zanderigo, Scott Houliston, Jennifer C Liddle, Chad M Dashnaw, T Michael Sabo, Bryan F Shaw, Jeremy L Balsbaugh, Gabriel J Rocklin, Colin A Smith
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引用次数: 0

摘要

最近出现了一种新技术,即从头设计微型蛋白抑制剂,以制造能与特定治疗靶点高亲和力结合的蛋白质。它们的大小、易表达性和明显的高稳定性使它们成为一类新型蛋白质药物的极佳候选者。然而,除了圆二色性熔融和氢/氘交换实验之外,人们对它们的动力学知之甚少,尤其是在它们似乎能很好耐受的高温条件下。为了解决这个问题并为未来的设计提供洞察力,我们重点研究了一种特别稳定的模型小蛋白 EHEE_rd2_0005,它的结构和化学变化是由热引起的,而这些变化以前被忽视了。核磁共振(NMR)研究表明,在多个时间和温度尺度上都存在动态变化。瞬时升温会导致核磁共振光谱中可见的自发化学脱酰胺作用,我们使用毛细管电泳和质谱(MS)实验对此进行了验证。高温还导致氢交换的内在速率大大加快,核磁共振异核单量子相干光谱中的信号因局部展开而损失。这些损失与室温氢交换实验和复制交换分子动力学模拟中的氢键破坏非常吻合。我们的分析揭示了未来微型蛋白设计的重要原则,以及高稳定性导致长寿命交替构象态的潜力。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Heat-induced structural and chemical changes to a computationally designed miniprotein.

The de novo design of miniprotein inhibitors has recently emerged as a new technology to create proteins that bind with high affinity to specific therapeutic targets. Their size, ease of expression, and apparent high stability makes them excellent candidates for a new class of protein drugs. However, beyond circular dichroism melts and hydrogen/deuterium exchange experiments, little is known about their dynamics, especially at the elevated temperatures they seemingly tolerate quite well. To address that and gain insight for future designs, we have focused on identifying unintended and previously overlooked heat-induced structural and chemical changes in a particularly stable model miniprotein, EHEE_rd2_0005. Nuclear magnetic resonance (NMR) studies suggest the presence of dynamics on multiple time and temperature scales. Transiently elevating the temperature results in spontaneous chemical deamidation visible in the NMR spectra, which we validate using both capillary electrophoresis and mass spectrometry (MS) experiments. High temperatures also result in greatly accelerated intrinsic rates of hydrogen exchange and signal loss in NMR heteronuclear single quantum coherence spectra from local unfolding. These losses are in excellent agreement with both room temperature hydrogen exchange experiments and hydrogen bond disruption in replica exchange molecular dynamics simulations. Our analysis reveals important principles for future miniprotein designs and the potential for high stability to result in long-lived alternate conformational states.

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来源期刊
Protein Science
Protein Science 生物-生化与分子生物学
CiteScore
12.40
自引率
1.20%
发文量
246
审稿时长
1 months
期刊介绍: Protein Science, the flagship journal of The Protein Society, is a publication that focuses on advancing fundamental knowledge in the field of protein molecules. The journal welcomes original reports and review articles that contribute to our understanding of protein function, structure, folding, design, and evolution. Additionally, Protein Science encourages papers that explore the applications of protein science in various areas such as therapeutics, protein-based biomaterials, bionanotechnology, synthetic biology, and bioelectronics. The journal accepts manuscript submissions in any suitable format for review, with the requirement of converting the manuscript to journal-style format only upon acceptance for publication. Protein Science is indexed and abstracted in numerous databases, including the Agricultural & Environmental Science Database (ProQuest), Biological Science Database (ProQuest), CAS: Chemical Abstracts Service (ACS), Embase (Elsevier), Health & Medical Collection (ProQuest), Health Research Premium Collection (ProQuest), Materials Science & Engineering Database (ProQuest), MEDLINE/PubMed (NLM), Natural Science Collection (ProQuest), and SciTech Premium Collection (ProQuest).
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