Variants of the Acyltransferase fromMycobacterium smegmatisEnable Enantioselective Acyl Transfer in Water

Abstract

The acyltransferase from Mycobacterium smegmatis (MsAcT) complements the well-established acylation activity of hydrolases in organic solvents with its activity to perform acylation reactions (among other reactions) in an aqueous environment. The enzyme’s potential is however limited, due to its poor regio- and stereoselectivity with enantioselectivities (E-values) below 20 for bulky (aromatic) substrates. By applying computer-guided rational design, a library of single variants was designed that allowed conversion of a set of previously challenging substrates with good activity and E-values up to >200. The computational predictions were found to be in agreement with experimental data, which in turn allowed for the generation of even more active and selective double variants. Overall, the produced set of variants provides a toolbox for the enantioselective acylation of challenging alcohols in water, effectively contributing to an alternative to reactions in organic solvents.

Discovery and Design of Family VIII Carboxylesterases as Highly Efficient Acyltransferases

“…Moreover, they can be used in cascade reactions with other enzymes that are not active or stable in the presence of organic solvents [4] . Since being reported in 2007, the promiscuous acyltransferase MsAcT from Mycobacterium smegmatis had been unrivalled in terms of transfer efficiency [5–7] . However, we recently demonstrated that many esterases from the bacterial hormone‐sensitive lipase (bHSL) family have promiscuous acyltransferase activity, some comparable to that of MsAcT, suggesting that this phenomenon might be more widespread than previously thought [8] .…”

“…[4] Since being reported in 2007, the promiscuous acyltransferase MsAcT from Mycobacterium smegmatis had been unrivalled in terms of transfer efficiency. [5][6][7] However, we recently demonstrated that many esterases from the bacterial hormone-sensitive lipase (bHSL) family have promiscuous acyltransferase activity, some comparable to that of MsAcT, suggesting that this phenomenon might be more widespread than previously thought. [8] However, enzymatic product hydrolysis and low transfer efficiency has always been a major drawback for application on industrial scale.…”

Power of Biocatalysis for Organic Synthesis

“… 133 The toolbox of available enzymes of the class is readily growing, with the most recent addition being acyltransferases that are able to catalyze transesterifications even in an aqueous environment. 134 − 136 …”

Entdeckung und Design promiskuitiver Acyltransferase‐Aktivität in Carboxylesterasen der Familie VIII

“…Dies ermöglicht außerdem den Einsatz in Kaskadenreaktionen zusammen mit Enzymen, die in Gegenwart organischer Lösungsmittel inaktiv oder instabil sind [4] . Seit ihrer Entdeckung im Jahr 2007 war die promiskuitive Acyltransferase MsAcT aus Mycobacterium smegmatis hinsichtlich der Effizienz des Acyltransfers in Wasser unangefochten [5–7] . Kürzlich konnten wir jedoch zeigen, dass viele Esterasen aus der Familie der bakteriellen hormonsensitiven Lipasen (bHSL) ebenfalls promiskuitive Acyltransferase‐Aktivität aufweisen [8] .…”

“…[4] Seit ihrer Entdeckung im Jahr 2007 war die promiskuitive Acyltransferase MsAcT aus Mycobacterium smegmatis hinsichtlich der Effizienz des Acyltransfers in Wasser unangefochten. [5][6][7] Kürzlich konnten wir jedoch zeigen, dass viele Esterasen aus der Familie der bakteriellen hormonsensitiven Lipasen (bHSL) ebenfalls promiskuitive Acyltransferase-Aktivität aufweisen. [8] Manche dieser Enzyme sind hinsichtlich ihrer Effizienz vergleichbar mit MsAcT, was darauf hindeutet, dass dieses Phänomen weiter verbreitet ist, als bisher angenommen.…”