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.
Keywords: Catechol oxidase; Polyphenol oxidase; Tyrosinase; enzyme mechanism; substrate specificity.