Enantioselective lipase-catalyzed O-acylation of (RS)-propranolol: analysis of the hydrogen bonds essential for catalysis

We investigated the effect of the acyl group size in the enantioselectivity of the acylation of propranolol, an amino alcohol used as β-adrenergic blocking agent. We applied a methodology frequently used to model enantioselectivity that is based on the hydrogen bonds present in the tetrahedral intermediate, which occurs in lipase-catalyzed reactions. We sampled the conformations of the tetrahedral intermediate corresponding to the esterification of both enantiomers of propranolol with ethanoyl and butanoyl, employing molecular dynamics simulation together with a quantum mechanics/molecular mechanics approach. We found that the population of these hydrogen bonds provides insight into the mechanism of the reaction. However, they are not conclusive about the role of the acyl group in the enantioselectivity. For both acyl groups, we found that the reaction from the Michaelis complex to the tetrahedral intermediate is more favorable for (R)-propranolol and the reaction from the tetrahedral intermediate to the enzyme/product complex is more favorable for (S)-propranolol.

Crystal structures of γ-glutamylmethylamide synthetase provide insight into bacterial metabolism of oceanic monomethylamine

Monomethylamine (MMA) is an important climate-active oceanic trace gas and ubiquitous in the oceans. The γ-glutamylmethylamide synthetase (GmaS) catalyzes the conversion of MMA to γ-glutamylmethylamide (GMA), the first step in MMA metabolism in many marine bacteria. The gmaS gene occurs in ~23% of microbial genomes in the surface ocean and is a validated biomarker to detect MMA-utilizing bacteria. However, the catalytic mechanism of GmaS has not been studied due to the lack of structural information. Here, the GmaS from Rhodovulum sp. 12E13 (RhGmaS) was characterized, and the crystal structures of apo-RhGmaS and RhGmaS with different ligands in five states were solved. Based on structural and biochemical analyses, the catalytic mechanism of RhGmaS was explained. ATP is first bound in RhGmaS, leading to a conformational change of a flexible loop (Lys287-Ile305), which is essential for the subsequent binding of glutamate. During the catalysis of RhGmaS, the residue Arg312 participates in polarizing the γ-phosphate of ATP and in stabilizing the γ-glutamyl phosphate intermediate; Asp177 is responsible for the deprotonation of MMA, assisting the attack of MMA on γ-glutamyl phosphate to produce a tetrahedral intermediate; and Glu186 acts as a catalytic base to abstract a proton from the tetrahedral intermediate to finally generate GMA. Sequence analysis suggested that the catalytic mechanism of RhGmaS proposed in this study has universal significance in bacteria containing GmaS. Our results provide novel insights into MMA metabolism, contributing to a better understanding of MMA catabolism in global carbon and nitrogen cycles.

Thioesterification and Selenoesterification of Amides via Selective N–C Cleavage at Room Temperature: N–C(O) to S/Se–C(O) Interconversion

The direct nucleophilic addition to amides represents an attractive methodology in organic synthesis that tackles amidic resonance by ground-state destabilization. This approach has been recently accomplished with carbon, nitrogen and oxygen nucleophiles. Herein, we report an exceedingly mild method for the direct thioesterification and selenoesterification of amides by selective N–C(O) bond cleavage in the absence of transition metals. Acyclic amides undergo N–C(O) to S/Se–C(O) interconversion to give the corresponding thioesters and selenoesters in excellent yields at room temperature via a tetrahedral intermediate pathway (cf. an acyl metal).

Phosphate-Catalyzed Succinimide Formation from an NGR-Containing Cyclic Peptide: A Novel Mechanism for Deammoniation of the Tetrahedral Intermediate

Spontaneous deamidation in the Asn-Gly-Arg (NGR) motif that yields an isoAsp-Gly-Arg (isoDGR) sequence has recently attracted considerable attention because of the possibility of application to dual tumor targeting. It is well known that Asn deamidation reactions in peptide chains occur via the five-membered ring succinimide intermediate. Recently, we computationally showed by the B3LYP density functional theory method, that inorganic phosphate and the Arg side chain can catalyze the NGR deamidation using a cyclic peptide, c[CH2CO–NGRC]–NH2. In this previous study, the tetrahedral intermediate of the succinimide formation was assumed to be readily protonated at the nitrogen originating from the Asn side chain by the solvent water before the release of an NH3 molecule. In the present study, we found a new mechanism for the decomposition of the tetrahedral intermediate that does not require the protonation by an external proton source. The computational method is the same as in the previous study. In the new mechanism, the release of an NH3 molecule occurs after a proton exchange between the peptide and the phosphate and conformational changes. The rate-determining step of the overall reaction course is the previously reported first step, i.e., the cyclization to form the tetrahedral intermediate.

1,3,4-Oxadiazole and Heteroaromatic-Fused 1,2,4-Triazole Synthesis­ Using Diverted Umpolung Amide Synthesis

Umpolung Amide Synthesis (UmAS) has emerged as a superior alternative to conventional amide synthesis methods based on carbonyl electrophiles in a range of situations, particularly when epimerization-prone couplings are prescribed. In an unanticipated development during our most recent studies, we discovered that diacyl hydrazide products from UmAS were not formed as intermediates when using an acyl hydrazide as the amine acceptor. This resulted in a new preparation of 1,3,4-oxadiazoles from α-bromonitroalkane donors. We hypothesized that a key tetrahedral intermediate in UmAS was diverted toward a more direct pathway to the heterocycle product rather than through formation of the diacyl hydrazide, a typical oxadiazole progenitor. In studies reported here, diversion to 1,2,4-triazole products is described, a behavior hypothesized to also result from an analogous tetrahedral intermediate, but one formed from heteroaromatic hydrazine acceptors.