The ubiquitous type-3 copper enzyme polyphenol oxidase (PPO) has found itself the subject of profound inhibitor research due to its role in fruit and vegetable browning and mammalian pigmentation. The enzyme itself has also been applied in the fields of bioremediation, biocatalysis and biosensing. However, the nature of PPO substrate specificity has remained elusive despite years of study. Numerous theories have been proposed to account for the difference in tyrosinase and catechol oxidase activity. The “blocker residue” theory suggests that bulky residues near the active site cover CuA, preventing monophenol coordination. The “second shell” theory suggests that residues distant (∼8 Å) from the active site, guide and position substrates within the active site based on their properties e.g., hydrophobic, electrostatic. It is also hypothesized that binding specificity is related to oxidation mechanisms of the catalytic cycle, conferred by coordination of a conserved water molecule by other conserved residues. In this review, we highlight recent developments in the structural and mechanistic studies of PPOs and consolidate key concepts in our understanding toward the substrate specificity of PPOs.

普遍存在的3型铜酶多酚氧化酶（PPO）由于其在水果和蔬菜褐变和哺乳动物色素沉着中的作用而已成为广泛的抑制剂研究对象。该酶本身也已应用于生物修复，生物催化和生物传感领域。然而，尽管经过多年的研究，PPO底物特异性的本质仍然难以捉摸。已经提出了许多理论来解释酪氨酸酶和邻苯二酚氧化酶活性的差异。“阻滞剂残基”理论表明，活性位点附近的庞大残基覆盖了CuA，从而阻止了单酚的配位。“第二壳”理论表明，与活性位点相距较远（约8埃）的残基会根据其特性（例如疏水性，静电性）在活性位点内引导和定位底物。还假设结合特异性与催化循环的氧化机制有关，这是由其他保守残基对保守水分子的配位而赋予的。在这篇综述中，我们重点介绍了PPO的结构和机理研究的最新进展，并巩固了我们对PPO底物特异性的理解中的关键概念。