Constitutively solvent-tolerant Pseudomonas taiwanensis VLB120 increment C increment ttgV supports particularly high-styrene epoxidation activities when grown under glucose excess conditions

摘要

Solvent-tolerant bacteria represent an interesting option to deal with the substrate and product toxicity in bioprocesses. Recently, constitutive solvent tolerance was achieved for Pseudomonas taiwanensis VLB120 via knockout of the regulator TtgV, making tedious adaptation unnecessary. Remarkably, ttgV knockout increased styrene epoxidation activities of P. taiwanensis VLB120 Delta C. With the aim to characterize and exploit the biocatalytic potential of P. taiwanensis VLB120 Delta C and VLB120 Delta C Delta ttgV, we investigated and correlated growth physiology, native styrene monooxygenase (StyAB) gene expression, whole-cell bioconversion kinetics, and epoxidation performance. Substrate inhibition kinetics was identified but was attenuated in two-liquid phase bioreactor setups. StyA fusion to the enhanced green fluorescent protein enabled precise enzyme level monitoring without affecting epoxidation activity. Glucose limitation compromised styAB expression and specific activities (30-40 U/g(CDW) for both strains), whereas unlimited batch cultivation enabled specific activities up to 180 U/g(CDW) for VLB120 Delta C Delta ttgV strains, which is unrivaled for bioreactor-based whole-cell oxygenase biocatalysis. These extraordinarily high specific activities of constitutively solvent-tolerant P. taiwanensis VLB120 increment C increment ttgV could be attributed to its high metabolic capacity, which also enabled high expression levels. This, together with the high product yields on glucose and biomass obtained qualifies the VLB120 increment ttgV strain as a highly attractive tool for the development of ecoefficient oxyfunctionalization processes and redox biocatalysis in general.