Heuschkel Ingeborg, Hanisch Selina, Volke Daniel C, Löfgren Erik, Hoschek Anna, Nikel Pablo I, Karande Rohan, Bühler Katja
Department of Solar Materials Helmholtz-Centre for Environmental Research Leipzig Germany.
ZINT - Zentrum für integrierte Naturstofftechnik TU Dresden Dresden Germany.
Eng Life Sci. 2021 Feb 2;21(3-4):258-269. doi: 10.1002/elsc.202000072. eCollection 2021 Mar.
In this study, the biocatalytic performance of a Baeyer-Villiger monooxygenase (BVMO) catalyzing the reaction of cyclohexanone to ε-caprolactone was investigated in biofilms. Biofilm growth and development of two VLB120 variants, Ps_BVMO and Ps_BVMO_DGC, were evaluated in drip flow reactors (DFRs) and rotating bed reactors (RBRs). Engineering a hyperactive diguanylate cyclase (DGC) from into Ps_BVMO resulted in faster biofilm growth compared to the control Ps_BVMO strain in the DFRs. The maximum product formation rates of 92 and 87 g m d were observed for mature Ps_BVMO and Ps_ BVMO_DGC biofilms, respectively. The application of the engineered variants in the RBR was challenged by low biofilm surface coverage (50-60%) of rotating bed cassettes, side-products formation, oxygen limitation, and a severe drop in production rates with time. By implementing an active oxygen supply mode and a twin capillary spray feed, the biofilm surface coverage was maximized to 70-80%. BVMO activity was severely inhibited by cyclohexanol formation, resulting in a decrease in product formation rates. By controlling the cyclohexanone feed concentration at 4 mM, a stable product formation rate of 14 g m d and a substrate conversion of 60% was achieved in the RBR.
在本研究中，对催化环己酮反应生成ε-己内酯的拜耳-维利格单加氧酶（BVMO）在生物膜中的生物催化性能进行了研究。在滴流反应器（DFR）和旋转床反应器（RBR）中评估了两种VLB120变体，即Ps_BVMO和Ps_BVMO_DGC的生物膜生长和发育情况。将一种来自[具体来源未提及]的高活性二鸟苷酸环化酶（DGC）工程化到Ps_BVMO中，与DFR中的对照Ps_BVMO菌株相比，导致生物膜生长更快。成熟的Ps_BVMO和Ps_BVMO_DGC生物膜的最大产物形成速率分别为92和87 g m⁻² d⁻¹。工程变体在RBR中的应用面临旋转床盒生物膜表面覆盖率低（50 - 60%）、副产物形成、氧气限制以及随着时间推移生产率严重下降等挑战。通过实施主动供氧模式和双毛细管喷雾进料，生物膜表面覆盖率最大化至70 - 80%。环己醇的形成严重抑制了BVMO活性，导致产物形成速率下降。通过将环己酮进料浓度控制在4 mM，在RBR中实现了14 g m⁻² d⁻¹的稳定产物形成速率和60%的底物转化率。
