Ester Synthesis in Water: Mycobacterium smegmatis Acyl Transferase for Kinetic Resolutions

Abstract

The acyl transferase from Mycobacterium smegmatis (MsAcT) catalyses transesterification reactions in aqueous media because of its hydrophobic active site. Aliphatic cyanohydrin and alkyne esters can be synthesised in water with excellent and strikingly opposite enantioselectivity [(R);E>37 and (S);E>100, respectively]. When using this enzyme, the undesired hydrolysis of the acyl donor is an important factor to take into account. Finally, the choice of acyl donor can significantly influence the obtained enantiomeric excesses.magnified image

Origins of Enantiopreference of Mycobacterium smegmatis Acyl Transferase: A Computational Analysis

“…Calculations predict the barrier for the S enantiomer to be 12.1 kcal mol −1 , relative to E‐Ac⋅ ( R )‐ 1 Up , whereas the barrier for the R enantiomer is calculated to be 17.1 kcal mol −1 , relative to the same starting structure. This result is in agreement with the experimental finding that MsAcT favors the S enantiomer of this substrate, with a measured E value of 63, corresponding to an energy difference of about 2 kcal mol −1 under the experimental conditions . The calculated energy difference of 5.0 kcal mol −1 is thus consistent with this, but somewhat overestimated, which could be due to the slight rigidity of the employed active‐site model.…”

“…Indeed, we have recalculated the two selectivity‐determining TSs for 1‐methyl propargyl alcohol ( 3 ; Scheme ) and the energy difference drops from 5.0 to 1.7 kcal mol −1 in favor of the S enantiomer (structures are given in the Supporting Information). This is in agreement with the measured E value for this substrate of only 15 (corresponding to an energy difference of about 1.4 kcal mol −1 ), compared with the E value of 63 for 1 …”

“…In this context, an enzyme that has great practical potential is the acyl transferase from Mycobacterium smegmatis (MsAcT), which is able to promote the transesterification reaction in aqueous solution . In addition, this enzyme is enantioselective for selected secondary alcohols, which makes it a potentially valuable tool in the production of optically active chemical compounds of interest for industrial applications. Similarly to lipases and esterases, the reaction of MsAcT involves two half‐reactions, in which the enzyme is acylated by an acyl donor in the first half‐reaction and the acyl group is then transferred to the substrate in the second half‐reaction.…”

“…Considering the potential of MsAcT for kinetic and dynamic resolution applications to be performed in aqueous medium instead of organic solvents, it is of great interest to provide a detailed understanding of MsAcT stereoselectivity. Propargylic alcohols and cyanohydrins are the most suitable substrates for this purpose because they exhibit excellent enantioselectivities, and, interestingly, the enzyme displays opposite enantiopreference for these substrates. In other words, MsAcT favors the R enantiomer of cyanohydrins, whereas, for propargylic alcohols, the S enantiomer is favored .…”

“…Propargylic alcohols and cyanohydrins are the most suitable substrates for this purpose because they exhibit excellent enantioselectivities, and, interestingly, the enzyme displays opposite enantiopreference for these substrates. In other words, MsAcT favors the R enantiomer of cyanohydrins, whereas, for propargylic alcohols, the S enantiomer is favored . This is quite puzzling, especially considering the structural similarities between these two substrate groups.…”