Bioconversion of renewable lignocellulosics to produce liquid fuels and chemicals is one of the most effective ways to solve the problem of fossil resource shortage, energy security, and environmental challenges. Among the many biorefinery pathways, hydrolysis of lignocellulosics to fermentable monosaccharides by cellulase is arguably the most critical step of lignocellulose bioconversion. In the process of enzymatic hydrolysis, the direct physical contact between enzymes and cellulose is an essential prerequisite for hydrolysis to occur. However, lignin is considered one of the most recalcitrant factors hindering the accessibility of cellulose by binding to cellulase unproductively, which reduces the saccharification rate and yield of carbohydrates. This results in high costs for the saccharification of carbohydrates. The various interactions between enzymes and lignin have been explored from different perspectives in literature, and a basic lignin inhibition mechanism has been proposed. However, the exact interaction between lignin and enzyme as well as the recently reported promotion of some types of lignin on enzymatic hydrolysis is still unclear at the molecular level. Multiple analytical techniques have been developed, and fully unlocking the secret of lignin-enzyme interactions would require a continuous improvement of the currently available techniques. This review summarizes the current commonly used advanced research analytical techniques for investigating the interaction between lignin and enzyme, including quartz crystal microbalance with dissipation (QCM-D), surface plasmon resonance (SPR), Fourier transform infrared spectroscopy-attenuated total reflectance (FTIR-ATR), atomic force microscopy (AFM), nuclear magnetic resonance (NMR) spectroscopy, fluorescence spectroscopy (FLS), and molecular dynamics (MD) simulations. Interdisciplinary integration of these analytical methods is pursued to provide new insight into the interactions between lignin and enzymes. This review will serve as a resource for future research seeking to develop new methodologies for a better understanding of the basic mechanism of lignin-enzyme binding during the critical hydrolysis process.

将可再生木质纤维素生物转化为液体燃料和化学品是解决化石资源短缺、能源安全和环境挑战的最有效方法之一。在众多生物精炼途径中，纤维素酶将木质纤维素水解为可发酵的单糖可以说是木质纤维素生物转化的最关键步骤。在酶水解过程中，酶与纤维素之间的直接物理接触是水解发生的必要前提。然而，木质素被认为是阻碍纤维素可及性的最顽固因素之一，它通过非生产性地与纤维素酶结合，从而降低了糖化率和碳水化合物的产量。这导致碳水化合物糖化的高成本。文献从不同角度探讨了酶与木质素之间的各种相互作用，并提出了一种基本的木质素抑制机制。然而，木质素与酶之间的确切相互作用以及最近报道的某些类型的木质素对酶水解的促进作用在分子水平上仍不清楚。已经开发了多种分析技术，完全解开木质素-酶相互作用的秘密需要不断改进当前可用的技术。本综述总结了目前研究木质素与酶相互作用的常用先进研究分析技术，包括带耗散的石英晶体微量天平（QCM-D）、表面等离子共振（SPR）、傅里叶变换红外光谱衰减全反射 (FTIR-ATR)、原子力显微镜 (AFM)、核磁共振 (NMR) 光谱、荧光光谱 (FLS) 和分子动力学 (MD) 模拟。追求这些分析方法的跨学科整合，以提供对木质素和酶之间相互作用的新见解。本综述将作为未来研究的资源，旨在开发新的方法，以更好地了解关键水解过程中木质素-酶结合的基本机制。追求这些分析方法的跨学科整合，以提供对木质素和酶之间相互作用的新见解。本综述将作为未来研究的资源，旨在开发新的方法，以更好地了解关键水解过程中木质素-酶结合的基本机制。追求这些分析方法的跨学科整合，以提供对木质素和酶之间相互作用的新见解。本综述将作为未来研究的资源，旨在开发新的方法，以更好地了解关键水解过程中木质素-酶结合的基本机制。