Natural production of anti-cancer drug taxol from Taxus has proved to be environmentally unsustainable and economically unfeasible. Currently, bioengineering the biosynthetic pathway of taxol is an attractive alternative production approach. 10-deacetylbaccatin III-10-O-acetyl transferase (DBAT) was previously characterized as an acyltransferase, using 10-deacetylbaccatin III (10-DAB) and acetyl CoA as natural substrates, to form baccatin III in the taxol biosynthesis. Here, we report that other than the natural acetyl CoA (Ac-CoA) substrate, DBAT can also utilize vinyl acetate (VA), which is commercially available at very low cost, acylate quickly and irreversibly, as acetyl donor in the acyl transfer reaction to produce baccatin III. Furthermore, mutants were prepared via a semi-rational design in this work. A double mutant, I43S/D390R was constructed to combine the positive effects of the different single mutations on catalytic activity, and its catalytic efficiency towards 10-DAB and VA was successfully improved by 3.30-fold, compared to that of wild-type DBAT, while 2.99-fold higher than the catalytic efficiency of WT DBAT towards 10-DAB and Ac-CoA. These findings can provide a promising economically and environmentally friendly method for exploring novel acyl donors to engineer natural product pathways.

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Taxoid 10β-O-acetyl transferase mutants with redesigned active sites displayed increased catalytic activities and modified substrate preferences, indicating their possible application in the enzymatic synthesis of baccatin III and taxol.

Summary Anisyl alcohol is an important spice in flavouring industries. However, its volatility makes it easy to degrade in high‐temperature pretreatment, thus limiting its wider application in tobacco industry and food reheating. To develop thermal stable flavour of this spice, a novel flavour precursor anisalcohol‐β‐galactoside was synthesised by glycosylation reaction, using anisyl alcohol and D‐lactose as substrates and recombinant β‐galactosidase as catalyst. The thermal decomposition process was determined by thermogravimetric analysis (TGA). TG curves revealed the maximum weight loss of anisyl alcohol reached 75% between 90 and 190 °C, while that of anisicohol‐β‐galactosidase was only 54% between 240 and 360 °C, which was an even higher range of temperature. The optimisation in thermal behaviour of anisicohol‐β‐galactosidase made it prospectively available in high‐temperature pretreatment and provided a new method for thermal stability improvement of other similar spices.