Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures

Seo, Hyeongmin; Lee, Jong-Won; Garcia, Sergio; Trinh, Cong T.

doi:10.1186/s13068-019-1583-8

Single mutation at a highly conserved region of chloramphenicol acetyltransferase enables isobutyl acetate production directly from cellulose by Clostridium thermocellum at elevated temperatures

Journal Article · Tue Oct 15 00:00:00 EDT 2019 · Biotechnology for Biofuels

DOI:https://doi.org/10.1186/s13068-019-1583-8· OSTI ID:1618766

Seo, Hyeongmin; Lee, Jong-Won; Garcia, Sergio;

Abstract Background

Esters are versatile chemicals and potential drop-in biofuels. To develop a sustainable production platform, microbial ester biosynthesis using alcohol acetyltransferases (AATs) has been studied for decades. Volatility of esters endows high-temperature fermentation with advantageous downstream product separation. However, due to the limited thermostability of AATs known, the ester biosynthesis has largely relied on use of mesophilic microbes. Therefore, developing thermostable AATs is important for ester production directly from lignocellulosic biomass by the thermophilic consolidated bioprocessing (CBP) microbes, e.g., Clostridium thermocellum .

Results

In this study, we engineered a thermostable chloramphenicol acetyltransferase from Staphylococcus aureus (CAT _Sa ) for enhanced isobutyl acetate production at elevated temperatures. We first analyzed the broad alcohol substrate range of CAT _Sa . Then, we targeted a highly conserved region in the binding pocket of CAT _Sa for mutagenesis. The mutagenesis revealed that F97W significantly increased conversion of isobutanol to isobutyl acetate. Using CAT _Sa F97W, we demonstrated direct conversion of cellulose into isobutyl acetate by an engineered C. thermocellum at elevated temperatures.

Conclusions

This study highlights that CAT is a potential thermostable AAT that can be harnessed to develop the thermophilic CBP microbial platform for biosynthesis of designer bioesters directly from lignocellulosic biomass.

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Research Organization:: Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)

Sponsoring Organization:: Center for Bioenergy Innovation; National Science Foundation; USDOE; USDOE Office of Science (SC)

Grant/Contract Number:: AC02-05CH11231

OSTI ID:: 1618766

Alternate ID(s):: OSTI ID: 1627005

Journal Information:: Biotechnology for Biofuels, Journal Name: Biotechnology for Biofuels Journal Issue: 1 Vol. 12; ISSN 1754-6834

Publisher:: Springer Science + Business MediaCopyright Statement

Country of Publication:: Netherlands

Language:: English

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