Engineering a hemoglobin variant with enhanced stability and oxygen affinity using a protein language model.

IF 5.3 3区化学 Q2 CHEMISTRY, MULTIDISCIPLINARY

Frontiers in Chemistry Pub Date : 2026-10-05 eCollection Date: 2026-01-01 DOI:10.3389/fchem.2026.1898272

Sergey Shityakov, Alexander S Novikov, Ekaterina V Skorb, Thomas Dandekar

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引用次数: 0

Abstract

Introduction: Engineered hemoglobin variants offer therapeutic promise for artificial blood substitutes and oxygen therapeutics, but rational design is constrained by protein stability and heme-binding. Protein language models (pLMs) predict mutation fitness, yet their integration with biophysical validation remains limited.

Methods: We developed a hybrid pipeline combining pLM (ESM3) with molecular docking, Monte Carlo simulation, molecular dynamics, MM-GBSA/PBSA binding free energy calculations, and density functional theory (DFT) to engineer a stabilized hemoglobin variant (esmHb) with enhanced oxygen affinity, benchmarking against the clinically characterized Hb Fort de France mutation (α45 His→Arg).

Results: ESM3 accurately assessed the fitness effect of the Hb Fort de France mutation and generated structurally stable esmHb models. Compared with the wild-type variant (wtHb), esmHb exhibited improved thermodynamic stability (ΔG: -18.48 vs. -12.22 kcal/mol), increased heme-binding affinity (ΔGbind: -11.79 vs. -8.42 kcal/mol), and greater thermodynamic favorability of oxidation according to DFT calculations (ΔE: -173.1 vs. -149.9 kcal/mol). MM-GBSA and MM-PBSA binding free energy calculations further supported improved protein-heme and protein-protein interactions, with esmHb showing more favorable values in both implicit solvation models. Statistical significance (p < 0.05) was reached for protein-protein interactions via both methods and for protein-heme interactions via MM-PBSA. We interpret the MM-GBSA results as a suggestive trend rather than definitive evidence.

Discussion: This work establishes a generalizable framework for pLM-driven protein engineering applicable to therapeutic Hb variants and beyond.

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利用蛋白质语言模型设计一种具有增强稳定性和氧亲和力的血红蛋白变体。

工程血红蛋白变体为人工血液替代品和氧气疗法提供了治疗前景，但合理的设计受到蛋白质稳定性和血红素结合的限制。蛋白质语言模型（pLMs）预测突变适应度，但其与生物物理验证的整合仍然有限。方法：我们开发了一种结合pLM （ESM3）与分子对接、蒙特卡罗模拟、分子动力学、MM-GBSA/PBSA结合自由能计算和密度泛函数理论（DFT）的混合管道，设计出具有增强氧亲和力的稳定血红蛋白变体（esmHb），以临床特征Hb Fort de France突变（α45 His→Arg）为基准。结果：ESM3准确评估了Hb Fort de France突变的适应度效应，生成了结构稳定的esmHb模型。与野生型变异（wtHb）相比，esmHb表现出更好的热力学稳定性（ΔG: -18.48 vs. -12.22 kcal/mol），增加血红素结合亲和力（ΔGbind: -11.79 vs. -8.42 kcal/mol），根据DFT计算（ΔE: -173.1 vs. -149.9 kcal/mol），更有利于氧化的热力学。MM-GBSA和MM-PBSA结合自由能计算进一步支持了蛋白质-血红素和蛋白质-蛋白质相互作用的改善，esmHb在两种隐式溶剂化模型中都显示出更有利的值。两种方法检测的蛋白相互作用和MM-PBSA检测的蛋白-血红素相互作用均达到统计学意义（p < 0.05）。我们将MM-GBSA结果解释为一种暗示性趋势，而不是明确的证据。讨论：这项工作为plm驱动的蛋白质工程建立了一个可推广的框架，适用于治疗性Hb变异及其他。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Frontiers in Chemistry Chemistry-General Chemistry

CiteScore

8.50

自引率

3.60%

发文量

1540

审稿时长

12 weeks

期刊介绍： Frontiers in Chemistry is a high visiblity and quality journal, publishing rigorously peer-reviewed research across the chemical sciences. Field Chief Editor Steve Suib at the University of Connecticut is supported by an outstanding Editorial Board of international researchers. This multidisciplinary open-access journal is at the forefront of disseminating and communicating scientific knowledge and impactful discoveries to academics, industry leaders and the public worldwide. Chemistry is a branch of science that is linked to all other main fields of research. The omnipresence of Chemistry is apparent in our everyday lives from the electronic devices that we all use to communicate, to foods we eat, to our health and well-being, to the different forms of energy that we use. While there are many subtopics and specialties of Chemistry, the fundamental link in all these areas is how atoms, ions, and molecules come together and come apart in what some have come to call the “dance of life”. All specialty sections of Frontiers in Chemistry are open-access with the goal of publishing outstanding research publications, review articles, commentaries, and ideas about various aspects of Chemistry. The past forms of publication often have specific subdisciplines, most commonly of analytical, inorganic, organic and physical chemistries, but these days those lines and boxes are quite blurry and the silos of those disciplines appear to be eroding. Chemistry is important to both fundamental and applied areas of research and manufacturing, and indeed the outlines of academic versus industrial research are also often artificial. Collaborative research across all specialty areas of Chemistry is highly encouraged and supported as we move forward. These are exciting times and the field of Chemistry is an important and significant contributor to our collective knowledge.