Whole-cell catalysis by surface display of fluorinase on Escherichia coli using N-terminal domain of ice nucleation protein

Abstract

Background Fluorinases play a unique role in the production of fluorine-containing organic molecules by biological methods. Whole-cell catalysis is a better choice in the large-scale fermentation processes, and over 60% of industrial biocatalysis uses this method. However, the in vivo catalytic efficiency of fluorinases is stuck with the mass transfer of the substrates. Results A gene sequence encoding a protein with fluorinase function was fused to the N-terminal of ice nucleation protein, and the fused fluorinase was expressed in Escherichia coli BL21(DE3) cells. SDS-PAGE and immunofluorescence microscopy were used to demonstrate the surface localization of the fusion protein. The fluorinase displayed on the surface showed good stability while retaining the catalytic activity. The engineered E.coli with surface-displayed fluorinase could be cultured to obtain a larger cell density, which was beneficial for industrial application. And 55% yield of 5′-fluorodeoxyadenosine (5′-FDA) from S-adenosyl-L-methionine (SAM) was achieved by using the whole-cell catalyst. Conclusions Here, we created the fluorinase-containing surface display system on E.coli cells for the first time. The fluorinase was successfully displayed on the surface of E.coli and maintained its catalytic activity. The surface display provides a new solution for the industrial application of biological fluorination. Graphical Abstract

Display of a novel carboxylesterase CarCby on Escherichia coli cell surface for carbaryl pesticide bioremediation

“…Surface-displayed strategies that allow peptides or target proteins to be located on the outer membrane of the cell surface have been widely used, such as biocatalyst, biodetoxification, biosensors, live vaccines, and peptide library screening for environmental, industrial, or diagnosis purposes [ 29 , 30 ]. Localization of target proteins on the cell surface could be used as a whole-cell biocatalyst directly because the substrates do not need to be transported across the membrane into the cells [ 6 ].…”

Biocatalytic synthesis of 2-fluoro-3-hydroxypropionic acid

“…On the one hand, fluoride ions have an inhibitory effect on the growth of E. coli ( Last et al, 2016 ). On the other hand, fluorinase requires the expensive co-substrate S-adenosyl methionine (SAM) ( Li et al, 2010 ; Feng et al, 2021 ; Kittila et al, 2022 ), and transmembrane transport of SAM requires the assistance of transporters ( Schaffitzel et al, 1998 ). These all limit the catalytic efficiency of fluorinase.…”