Lignin is one of the world's most abundant organic polymers, and 2-pyrone-4,6-dicarboxylate lactonase (LigI) catalyzes the hydrolysis of 2-pyrone-4,6-dicarboxylate (PDC) in the degradation of lignin. The pH has profound effects on enzyme catalysis and therefore we studied this in the context of LigI. We found that changes of the pH mostly affects surface residues, while the residues at the active site are more subject to changes of the surrounding microenvironment. In accordance with this, a high pH facilitates the deprotonation of the substrate. Detailed free energy calculations by the empirical valence bond (EVB) approach revealed that the overall hydrolysis reaction is more likely when the three active site histidines (His31, His33 and His180) are protonated at the ɛ site, however, protonation at the δ site may be favored during specific steps of the reaction. Our studies have uncovered the determinant role of the protonation state of the active site residues His31, His33 and His180 in the hydrolysis of PDC.

中文翻译：

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组氨酸质子化状态是 LigI 催化反应机制的关键

木质素是世界上最丰富的有机聚合物之一，2-pyrone-4,6-dicarboxylate lactonase (LigI) 在木质素降解中催化 2-pyrone-4,6-dicarboxylate (PDC) 的水解。pH 值对酶催化具有深远的影响，因此我们在 LigI 的背景下对此进行了研究。我们发现 pH 值的变化主要影响表面残留物，而活性位点的残留物更容易受到周围微环境的变化的影响。据此，高pH有利于底物的去质子化。通过经验价键 (EVB) 方法进行的详细自由能计算表明，当三个活性位点组氨酸（His31、His33 和 His180）在 ɛ 位质子化时，整体水解反应更有可能发生，然而，在δ位点可能在反应的特定步骤中受到青睐。我们的研究揭示了活性位点残基 His31、His33 和 His180 的质子化状态在 PDC 水解中的决定性作用。