Optimization of ribosomal binding site sequences for gene expression and 4-hydroxyisoleucine biosynthesis in recombinant corynebacterium glutamicum

Bacterial metabolism shifts from aerobic respiration to fermentation at the transition from exponential to stationary growth phases in response to limited oxygen availability. Corynebacterium glutamicum, a Gram-positive, facultative aerobic bacterium used for industrial amino acid production, excretes L-lactate, acetate, and succinate as fermentation products. The ldhA gene encoding L-lactate dehydrogenase is solely responsible for L-lactate production. Its expression is repressed at the exponential phase and prominently induced at the transition phase. ldhA is transcriptionally repressed by the sugar-phosphate-responsive regulator SugR and L-lactate-responsive regulator LldR. Although ldhA expression is derepressed even at the exponential phase in the sugR and lldR double deletion mutant, a further increase in its expression is still observed at the stationary phase, implicating the action of additional transcription regulators. In this study, involvement of the cAMP receptor protein-type global regulator GlxR in the regulation of ldhA expression was investigated. The GlxR-binding site found in the ldhA promoter was modified to inhibit or enhance binding of GlxR. The ldhA promoter activity and expression of ldhA were altered in proportion to the binding affinity of GlxR. Similarly, L-lactate production was also affected by the binding site modification. Thus, GlxR was demonstrated to act as a transcriptional activator of ldhA.

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Evidence that the requirements for ATP and wheat germ initiation factors 4A and 4F are affected by a region of satellite tobacco necrosis virus RNA that is 3' to the ribosomal binding site.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)68488-1 ◽

1988 ◽

Vol 263 (17) ◽

pp. 8380-8383 ◽

Cited By ~ 8

Author(s):

K S Browning ◽

L Fletcher ◽

J M Ravel

Keyword(s):

Binding Site ◽

Wheat Germ ◽

Tobacco Necrosis Virus ◽

Necrosis Virus ◽

Initiation Factors ◽

Virus Rna ◽

Ribosomal Binding Site

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Hesx1 homeodomain protein represses transcription as a monomer and antagonises transactivation of specific sites as a homodimer

Journal of Molecular Endocrinology ◽

10.1677/jme.0.0280193 ◽

2002 ◽

Vol 28 (3) ◽

pp. 193-205 ◽

Cited By ~ 10

Author(s):

J Quirk ◽

P Brown

Keyword(s):

Gene Expression ◽

Dna Binding ◽

Binding Site ◽

Anterior Pituitary ◽

Homeodomain Protein ◽

Dependent Manner ◽

Gene Promoters ◽

Differentiation Markers ◽

Proximal Half ◽

Dna Binding Site

The homeobox repressor Hesx1, expressed throughout Rathke's pouch and required for normal pituitary development, has been implicated in anterior pituitary pathogenesis in man. Prolonged expression of Hesx1 delays the appearance of anterior pituitary terminal differentiation markers in mice, particularly the gonadotroph hormones. We tested if Hesx1 could modulate gonadotrophin gene expression directly, and found that Hesx1 repressed both common alpha subunit (alpha GSU) and luteinising hormone beta-subunit (LH beta) gene promoters. Repression mapped to the Pitx1 homeodomain protein transactivation site in the proximal alpha GSU promoter, but did not map to the equivalent site on LH beta. Hesx1 repression of the alpha GSU Pitx1 site was overridden by co-transfection of Pitx1. In contrast, Hesx1 antagonised Pitx1 transactivation of LH beta in a dose-dependent manner. This was due to monomeric binding of Hesx1 on alpha GSU and homodimerisation on LH beta. The homodimerisation site comprises the Pitx1 DNA binding site and a proximal binding site, and mutation of either inhibited homodimer formation. Conversion of the LH beta Pitx1 DNA binding site to an alpha GSU-type did not promote homodimer formation, arguing that Hesx1 has pronounced site selectivity. Furthermore, mutation of the proximal half of the homodimerisation site blocked Hesx1 antagonisation of Pitx1 transactivation. We conclude that Hesx1 monomers repress gene expression, and homodimers block specific transactivation sites.

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Removal of Anionic Metalloid/Metals by PEI-Modified Corynebacterium glutamicum

Advanced Materials Research ◽

10.4028/www.scientific.net/amr.825.568 ◽

2013 ◽

Vol 825 ◽

pp. 568-571

Author(s):

Namgyu Kim ◽

Munsik Park ◽

Jongmoon Park ◽

Donghee Park

Keyword(s):

Aqueous Solution ◽

Binding Site ◽

Corynebacterium Glutamicum ◽

Adsorption Mechanism ◽

Competitive Inhibition ◽

Aqueous System ◽

Reduction Mechanism ◽

Removal Mechanism ◽

Sulfate Ion ◽

Industrial Fermentation

An anionic biosorbent was derived from an industrial fermentation biowate, Corynebacterium glutamicum, by being cross-linked with polyethylenimine (PEI). A fiber form of the biosorbent was used to examine its potentiality of removing anionic metals such as As (V), Cr (VI) and Mn (VII) in aqueous system. As (V) and Cr (VI) were efficiently removed by the biosorbent through anionic adsorption mechanism. Sulfate ion seriously inhibited adsorption of the anionic metals through competitive inhibition with respect to the binding site of the biosorbent. In the case of Mn (VII), its removal mechanism by the biosorbent was not anionic adsorption. Mn (VII) was completely removed in aqueous phase, meanwhile, Mn (II) appeared and increased in proportion to the Mn (VII) depletion. As a result, adsorption coupled reduction was chosen as the mechanism of Mn (VII) removal by the biosorbent. In conclusion, the anionic biosorbent could be used to remove various anionic metals from aqueous solution through anionic adsorption or reduction mechanism.

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Transforming growth factor-beta stimulates alpha 2(I) collagen gene expression through a cis-acting element that contains an Sp1-binding site.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(17)36699-1 ◽

1994 ◽

Vol 269 (20) ◽

pp. 14828-14834 ◽

Cited By ~ 2

Author(s):

Y. Inagaki ◽

S. Truter ◽

F. Ramirez

Keyword(s):

Gene Expression ◽

Growth Factor ◽

Binding Site ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Collagen Gene ◽

Cis Acting ◽

Collagen Gene Expression ◽

Growth Factor Beta

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Synergistic activation of ADH2 expression is sensitive to upstream activation sequence 2 (UAS2) orientation, copy number and UAS1-UAS2 helical phasing.

Molecular and Cellular Biology ◽

10.1128/mcb.15.6.3442 ◽

1995 ◽

Vol 15 (6) ◽

pp. 3442-3449 ◽

Cited By ~ 16

Author(s):

M S Donoviel ◽

N Kacherovsky ◽

E T Young

Keyword(s):

Gene Expression ◽

Binding Site ◽

Reporter Gene ◽

Copy Number ◽

Glucose Repression ◽

Regulatory Protein ◽

Regulatory Region ◽

Upstream Regulatory Region ◽

Specific Sequence

The alcohol dehydrogenase 2 (ADH2) gene of Saccharomyces cerevisiae is under stringent glucose repression. Two cis-acting upstream activation sequences (UAS) that function synergistically in the derepression of ADH2 gene expression have been identified. UAS1 is the binding site for the transcriptional regulator Adr1p. UAS2 has been shown to be important for ADH2 expression and confers glucose-regulated, ADR1-independent activity to a heterologous reporter gene. An analysis of point mutations within UAS2, in the context of the entire ADH2 upstream regulatory region, showed that the specific sequence of UAS2 is important for efficient derepression of ADH2, as would be expected if UAS2 were the binding site for a transcriptional regulatory protein. In the context of the ADH2 upstream regulatory region, including UAS1, working in concert with the ADH2 basal promoter elements, UAS2-dependent gene activation was dependent on orientation, copy number, and helix phase. Multimerization of UAS2, or its presence in reversed orientation, resulted in a decrease in ADH2 expression. In contrast, UAS2-dependent expression of a reporter gene containing the ADH2 basal promoter and coding sequence was enhanced by multimerization of UAS2 and was independent of UAS2 orientation. The reduced expression caused by multimerization of UAS2 in the native promoter was observed only in the presence of ADR1. Inhibition of UAS2-dependent gene expression by Adr1p was also observed with a UAS2-dependent ADH2 reporter gene. This inhibition increased with ADR1 copy number and required the DNA-binding activity of Adr1p. Specific but low-affinity binding of Adr1p to UAS2 in vitro was demonstrated, suggesting that the inhibition of UAS2-dependent gene expression observed in vivo could be a direct effect due to Adr1p binding to UAS2.

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