Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicum
Abstract Background Promoters are commonly used to regulate the expression of specific target genes or operons. Although a series of promoters have been developed in Corynebacterium glutamicum, more precise and unique expression patterns are needed that the current selection of promoters cannot prod...
Main Authors: | , , , , , |
---|---|
Format: | Article |
Language: | English |
Published: |
BMC
2018-11-01
|
Series: | Microbial Cell Factories |
Subjects: | |
Online Access: | http://link.springer.com/article/10.1186/s12934-018-1031-7 |
id |
doaj-6f2ddc8f69b048edb972add321730367 |
---|---|
record_format |
Article |
spelling |
doaj-6f2ddc8f69b048edb972add3217303672020-11-25T02:40:32ZengBMCMicrobial Cell Factories1475-28592018-11-0117111010.1186/s12934-018-1031-7Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicumYuechao Ma0Yi Cui1Lihong Du2Xiaoqian Liu3Xixian Xie4Ning Chen5National and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science & TechnologyNational and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science & TechnologyNational and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science & TechnologyNational and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science & TechnologyNational and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science & TechnologyNational and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science & TechnologyAbstract Background Promoters are commonly used to regulate the expression of specific target genes or operons. Although a series of promoters have been developed in Corynebacterium glutamicum, more precise and unique expression patterns are needed that the current selection of promoters cannot produce. RNA-Seq technology is a powerful tool for helping us to screen out promoters with expected transcriptional strengths. Results The promoter PCP_2836 of an aldehyde dehydrogenase coding gene from Corynebacterium glutamicum CP was identified via RNA-seq and RT-PCR as a growth-regulated promoter. Comparing with the strong constitutive promoter Ptuf, the transcriptional strength of PCP_2836 showed a significant decrease that from about 75 to 8% in the stationary phase. By replacing the native promoters of the aceE and gltA genes with PCP_2836 in the C. glutamicum ATCC 13032-derived l-valine-producing strain AN02, the relative transcriptional levels of the aceE and gltA genes decreased from 1.2 and 1.1 to 0.35 and 0.3, and the activity of their translation products decreased to 43% and 35%, respectively. After 28 h flask fermentation, the final cell density of the obtained strains, GRaceE and GRgltA, exhibited a 7–10% decrease. However, l-valine production increased by 23.9% and 27.3%, and the yield of substrate to product increased 43.8% and 62.5%, respectively. In addition, in the stationary phase, the intracellular citrate levels in GRaceE and GRgltA decreased to 27.0% and 33.6% of AN02, and their intracellular oxaloacetate levels increased to 2.7 and 3.0 times that of AN02, respectively. Conclusions The PCP_2836 promoter displayed a significant difference on its transcriptional strength in different cell growth phases. With using PCP_2836 to replace the native promoters of aceE and gltA genes, both the transcriptional levels of the aceE and gltA genes and the activity of their translation products demonstrated a significant decrease in the stationary phase. Thus, the availability of pyruvate was significantly increased for the synthesis of l-valine without any apparent irreversible negative impacts on cell growth. Use of this promoter can enhance the selectivity and control of gene expression and could serve as a useful research tool for metabolic engineering.http://link.springer.com/article/10.1186/s12934-018-1031-7Corynebacterium glutamicumGrowth-regulated promoterTranscriptional regulationPyruvate dehydrogenaseCitrate synthasel-Valine |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Yuechao Ma Yi Cui Lihong Du Xiaoqian Liu Xixian Xie Ning Chen |
spellingShingle |
Yuechao Ma Yi Cui Lihong Du Xiaoqian Liu Xixian Xie Ning Chen Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicum Microbial Cell Factories Corynebacterium glutamicum Growth-regulated promoter Transcriptional regulation Pyruvate dehydrogenase Citrate synthase l-Valine |
author_facet |
Yuechao Ma Yi Cui Lihong Du Xiaoqian Liu Xixian Xie Ning Chen |
author_sort |
Yuechao Ma |
title |
Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicum |
title_short |
Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicum |
title_full |
Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicum |
title_fullStr |
Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicum |
title_full_unstemmed |
Identification and application of a growth-regulated promoter for improving l-valine production in Corynebacterium glutamicum |
title_sort |
identification and application of a growth-regulated promoter for improving l-valine production in corynebacterium glutamicum |
publisher |
BMC |
series |
Microbial Cell Factories |
issn |
1475-2859 |
publishDate |
2018-11-01 |
description |
Abstract Background Promoters are commonly used to regulate the expression of specific target genes or operons. Although a series of promoters have been developed in Corynebacterium glutamicum, more precise and unique expression patterns are needed that the current selection of promoters cannot produce. RNA-Seq technology is a powerful tool for helping us to screen out promoters with expected transcriptional strengths. Results The promoter PCP_2836 of an aldehyde dehydrogenase coding gene from Corynebacterium glutamicum CP was identified via RNA-seq and RT-PCR as a growth-regulated promoter. Comparing with the strong constitutive promoter Ptuf, the transcriptional strength of PCP_2836 showed a significant decrease that from about 75 to 8% in the stationary phase. By replacing the native promoters of the aceE and gltA genes with PCP_2836 in the C. glutamicum ATCC 13032-derived l-valine-producing strain AN02, the relative transcriptional levels of the aceE and gltA genes decreased from 1.2 and 1.1 to 0.35 and 0.3, and the activity of their translation products decreased to 43% and 35%, respectively. After 28 h flask fermentation, the final cell density of the obtained strains, GRaceE and GRgltA, exhibited a 7–10% decrease. However, l-valine production increased by 23.9% and 27.3%, and the yield of substrate to product increased 43.8% and 62.5%, respectively. In addition, in the stationary phase, the intracellular citrate levels in GRaceE and GRgltA decreased to 27.0% and 33.6% of AN02, and their intracellular oxaloacetate levels increased to 2.7 and 3.0 times that of AN02, respectively. Conclusions The PCP_2836 promoter displayed a significant difference on its transcriptional strength in different cell growth phases. With using PCP_2836 to replace the native promoters of aceE and gltA genes, both the transcriptional levels of the aceE and gltA genes and the activity of their translation products demonstrated a significant decrease in the stationary phase. Thus, the availability of pyruvate was significantly increased for the synthesis of l-valine without any apparent irreversible negative impacts on cell growth. Use of this promoter can enhance the selectivity and control of gene expression and could serve as a useful research tool for metabolic engineering. |
topic |
Corynebacterium glutamicum Growth-regulated promoter Transcriptional regulation Pyruvate dehydrogenase Citrate synthase l-Valine |
url |
http://link.springer.com/article/10.1186/s12934-018-1031-7 |
work_keys_str_mv |
AT yuechaoma identificationandapplicationofagrowthregulatedpromoterforimprovinglvalineproductionincorynebacteriumglutamicum AT yicui identificationandapplicationofagrowthregulatedpromoterforimprovinglvalineproductionincorynebacteriumglutamicum AT lihongdu identificationandapplicationofagrowthregulatedpromoterforimprovinglvalineproductionincorynebacteriumglutamicum AT xiaoqianliu identificationandapplicationofagrowthregulatedpromoterforimprovinglvalineproductionincorynebacteriumglutamicum AT xixianxie identificationandapplicationofagrowthregulatedpromoterforimprovinglvalineproductionincorynebacteriumglutamicum AT ningchen identificationandapplicationofagrowthregulatedpromoterforimprovinglvalineproductionincorynebacteriumglutamicum |
_version_ |
1724781008726786048 |