“Bridge regions” regulate catalysis and protein stability of acylpeptide hydrolase

Abstract

Mutation in the enzyme engineering is a powerful tool that could alter enzyme properties, providing evidence for enzymology. Acylpeptide hydrolase (APH) is high research value enzyme which plays a key role in the scavenging of cytotoxic proteins and demonstrated a super efficiency in catalyzing CC bond formation in synthetic applications. Therefore, investigation of the structural and enzymatic properties is necessary. Herein, the results revealed that linker region that directly connects two domains of the monomer involves in stability and catalysis, and N-terminus is critical in mediating temperature-dependent catalysis. Moreover, the N-terminus and the linker jointly serve as bridge regions, that both offer a win-win strategy for catalysis and stability as the result of evolutionary adaptation to the existing environment. Based on these results, a rational design for mutation was proposed to optimize stability and promiscuous catalysis in aldol addition. It is worth noting that the mutant ST-4A (formed by inserting four Ala residues between K331 and G332 at the linker of ST0779) showed better stability and catalytic properties than the wild type, which is suitable for chemical synthesis. This study highlights the important function of bridge regions of APH and the deep understanding of catalysis and structure, provides a new direction for optimized enzymatic transformation and shows the potential value of APH members for synthetic applications.