Affiliation:
1. Research Institute for Bioscience Products & Fine Chemicals, Ajinomoto Co., Inc., Kawasaki, Japan
2. Ajinomoto-Genetika Research Institute, Moscow, Russia
Abstract
ABSTRACT
Cysteine is a commercially important amino acid; however, it lacks an efficient fermentative production method. Due to its cytotoxicity, intracellular cysteine levels are stringently controlled via several regulatory modes. Managing its toxic effects as well as understanding and deregulating the complexities of regulation are crucial for establishing the fermentative production of cysteine. The regulatory modes include feedback inhibition of key metabolic enzymes, degradation, efflux pumps, and the transcriptional regulation of biosynthetic genes by a master cysteine regulator, CysB. These processes have been extensively studied using
Escherichia coli
for overproducing cysteine by fermentation. In this study, we genetically engineered
Pantoea ananatis
, an emerging host for the fermentative production of bio-based materials, to identify key factors required for cysteine production. According to this and our previous studies, we identified a major cysteine desulfhydrase gene,
ccdA
(formerly PAJ_0331), involved in cysteine degradation, and the cysteine efflux pump genes
cefA
and
cefB
(formerly PAJ_3026 and PAJ_p0018, respectively), which may be responsible for downregulating the intracellular cysteine level. Our findings revealed that
ccdA
deletion and
cefA
and
cefB
overexpression are crucial factors for establishing fermentative cysteine production in
P. ananatis
and for obtaining a higher cysteine yield when combined with genes in the cysteine biosynthetic pathway. To our knowledge, this is the first demonstration of cysteine production in
P. ananatis
, which has fundamental implications for establishing overproduction in this microbe.
IMPORTANCE
The efficient production of cysteine is a major challenge in the amino acid fermentation industry. In this study, we identified cysteine efflux pumps and degradation pathways as essential elements and genetically engineered
Pantoea ananatis
, an emerging host for the fermentative production of bio-based materials, to establish the fermentative production of cysteine. This study provides crucial insights into the design and construction of cysteine-producing strains, which may play central roles in realizing commercial basis production.
Publisher
American Society for Microbiology
Subject
Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology
Cited by
39 articles.
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