Insight into Structural Characteristics of Protein-Substrate Interaction in Pimaricin Thioesterase

Abstract

As a polyene antibiotic of great pharmaceutical significance, pimaricin has been extensively studied to enhance its productivity and effectiveness. In our previous studies, pre-reaction state (PRS) has been validated as one of the significant conformational categories before macrocyclization, and is critical to mutual recognition and catalytic preparation in thioesterase (TE)-catalyzed systems. In our study, molecular dynamics (MD) simulations were conducted on pimaricin TE-polyketide complex and PRS, as well as pre-organization state (POS), a molecular conformation possessing a pivotal intra-molecular hydrogen bond, were detected. Conformational transition between POS and PRS was observed in one of the simulations, and POS was calculated to be energetically more stable than PRS by 4.58 kcal/mol. The structural characteristics of PRS and POS-based hydrogen-bonding, and hydrophobic interactions were uncovered, and additional simulations were carried out to rationalize the functions of several key residues (Q29, M210, and R186). Binding energies, obtained from MM/PBSA calculations, were further decomposed to residues, in order to reveal their roles in product release. Our study advanced a comprehensive understanding of pimaricin TE-catalyzed macrocyclization from the perspectives of conformational change, protein-polyketide recognition, and product release, and provided potential residues for rational modification of pimaricin TE.

Structure-Guided Engineering of d-Carbamoylase Reveals a Key Loop at Substrate Entrance Tunnel

“…The Bürgi–Dunitz angle is well-known to be the product of overlap between lowest unoccupied molecular orbital (LUMO) of the carbonyl and the highest occupied molecular orbital (HOMO) of the nucleophile, strongly suggesting that the elevated catalytic efficiency of M4 may be attributed to the increased interaction between LUMO and HOMO. The proportion of complexes in PRS is regarded as the degree of reaction readiness, which was 14.6% for M4– 3a , or 3.17 times that of WT– 3a (4.6%), therefore suggesting that M4 has increased reaction readiness compared with WT.…”