Optimizing the biosynthesis of oxygenated and acetylated Taxol precursors in Saccharomyces cerevisiae using advanced bioprocessing strategies.

Taxadien-5α-hydroxylase and taxadien-5α-ol O-acetyltransferase catalyze the oxidation of taxadiene to taxadien-5α-ol and subsequent acetylation to taxadien-5α-yl-acetate in the biosynthesis of the blockbuster anticancer drug, paclitaxel (Taxol®). Despite decades of research, the promiscuous and multispecific CYP725A4 enzyme remains a major bottleneck in microbial biosynthetic pathway development. In this study, an interdisciplinary approach was applied for the construction and optimization of the early pathway in Saccharomyces cerevisiae, across a range of bioreactor scales. High-throughput microscale optimization enhanced total oxygenated taxane titer to 39.0 ± 5.7 mg/L and total taxane product titers were comparable at micro and minibioreactor scale at 95.4 ± 18.0 and 98.9 mg/L, respectively. The introduction of pH control successfully mitigated a reduction of oxygenated taxane production, enhancing the potential taxadien-5α-ol isomer titer to 19.2 mg/L, comparable with the 23.8 ± 3.7 mg/L achieved at microscale. A combination of bioprocess optimization and increased gas chromatography-mass spectrometry resolution at 1 L bioreactor scale facilitated taxadien-5α-yl-acetate detection with a final titer of 3.7 mg/L. Total oxygenated taxane titers were improved 2.7-fold at this scale to 78 mg/L, the highest reported titer in yeast. Critical parameters affecting the productivity of the engineered strain were identified across a range of scales, providing a foundation for the development of robust integrated bioprocess control systems.

Micropollutant sorption to membrane polymers: a review of mechanisms for estrogens.

Organic micropollutants such as estrogens occur in water in increasing quantities from predominantly anthropogenic sources. In water such micropollutants partition not only to surfaces such as membrane polymers but also to any other natural or treatment related surfaces. Such interactions are often observed as sorption in treatment processes and this phenomenon is exploited in activated carbon filtration, for example. Sorption is important for polymeric materials and this is used for the concentration of such micropollutants for analytical purposes in solid phase extraction. In membrane filtration the mechanism of micropollutant sorption is a relatively new discovery that was facilitated through new analytical techniques. This sorption plays an important role in micropollutant retention by membranes although mechanisms of interaction are to date not understood. This review is focused on sorption of estrogens on polymeric surfaces, specifically membrane polymers. Such sorption has been observed to a large extent with values of up to 1.2 ng/cm(2) measured. Sorption is dependent on the type of polymer, micropollutant characteristics, solution chemistry, membrane operating conditions as well as membrane morphology. Likely contributors to sorption are the surface roughness as well as the microporosity of such polymers. While retention-and/or reflection coefficient as well as solute to effective pore size ratio-controls the access of such micropollutants to the inner surface, pore size, porosity and thickness as well as morphology or shape of inner voids determines the available area for sorption. The interaction mechanisms are governed, most likely, by hydrophobic as well as solvation effects and interplay of molecular and supramolecular interactions such as hydrogen bonding, π-cation/anion interactions, π-π stacking, ion-dipole and dipole-dipole interactions, the extent of which is naturally dependent on micropollutant and polymer characteristics. Systematic investigations are required to identify and quantify both relative contributions and strength of such interactions and develop suitable surface characterisation tools. This is a difficult endeavour given the complexity of systems, the possibility of several interactions taking place simultaneously and the generally weaker forces involved.