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Crystal Structure of TetR Family Repressor AlkX from Dietzia sp. Strain DQ12-45-1b Implicated in Biodegradation of n-Alkanes.

ABSTRACT: n-Alkanes are ubiquitous in nature and are widely used by microorganisms as carbon sources. Alkane hydroxylation by alkane monooxygenases is a critical step in the aerobic biodegradation of n-alkanes, which plays important roles in natural alkane attenuation and is used in industrial and environmental applications. The alkane oxidation operon, alkW1-alkX, in the alkane-degrading strain Dietzia sp. strain DQ12-45-1b is negatively autoregulated by the TetR family repressor AlkX via a product positive feedback mechanism. To predict the gene regulation mechanism, we determined the 3.1-Å crystal structure of an AlkX homodimer in a non-DNA-bound state. The structure showed traceable long electron density deep inside a hydrophobic cavity of each monomer along the long axis of the helix bundle, and further gas chromatography-mass spectrometry analysis of AlkX revealed that it contained the Escherichia coli-derived long-chain fatty acid molecules as a ligand. Moreover, an unusual structural feature of AlkX is an extra helix, ?6', forming a lid-like structure with ?6 covering the inducer-binding pocket and occupying the space between the two symmetrical DNA-binding motifs in one dimer, indicating a distinct conformational transition mode in modulating DNA binding. Sequence alignment of AlkX homologs from Dietzia strains showed that the residues involved in DNA and inducer binding are highly conserved, suggesting that the regulation mechanisms of n-alkane hydroxylation are possibly a common characteristic of Dietzia strains.IMPORTANCE With n-alkanes being ubiquitous in nature, many bacteria from terrestrial and aquatic environments have evolved n-alkane oxidation functions. Alkane hydroxylation by alkane monooxygenases is a critical step in the aerobic biodegradation of n-alkanes, which plays important roles in natural alkane attenuation and petroleum-contaminating environment bioremediation. The gene regulation of the most common alkane hydroxylase, AlkB, has been studied widely in Gram-negative bacteria but has been less explored in Gram-positive bacteria. Our previous study showed that the TetR family regulator (TFR) AlkX negatively autoregulated the alkane oxidation operon, alkW1-alkX, in the Gram-positive strain Dietzia sp. strain DQ12-45-1b. Although TFRs are one of the most common transcriptional regulator families in bacteria, the TFR involved in n-alkane metabolism has been reported only recently. In this study, we determined the crystal structure of AlkX, which implies a distinct DNA/ligand binding mode. Our results shed light upon the regulation mechanism of the common alkane degradation process in nature.

SUBMITTER: Liang JL

PROVIDER: S-EPMC5648916 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Crystal Structure of TetR Family Repressor AlkX from Dietzia sp. Strain DQ12-45-1b Implicated in Biodegradation of n-Alkanes.

Liang Jie-Liang JL Gao Yuan Y He Zheng Z Nie Yong Y Wang Meng M JiangYang Jing-Hong JH Zhang Xuejun C XC Shu Wen-Sheng WS Wu Xiao-Lei XL

Applied and environmental microbiology 20171017 21

n-Alkanes are ubiquitous in nature and are widely used by microorganisms as carbon sources. Alkane hydroxylation by alkane monooxygenases is a critical step in the aerobic biodegradation of n-alkanes, which plays important roles in natural alkane attenuation and is used in industrial and environmental applications. The alkane oxidation operon, alkW1-alkX, in the alkane-degrading strain Dietzia sp. strain DQ12-45-1b is negatively autoregulated by the TetR family repres ...[more]

PMID: 28821550

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Project description:Biodegradation of alkanes by microbial communities is ubiquitous in nature. Interestingly, the microbial communities with high hydrocarbon-degrading performances are sometimes composed of not only hydrocarbon degraders but also nonconsumers, but the synergistic mechanisms remain unknown. Here, we found that two bacterial strains isolated from Chinese oil fields, Dietzia sp. strain DQ12-45-1b and Pseudomonas stutzeri SLG510A3-8, had a synergistic effect on hexadecane (C16 compound) biodegradation, even though P. stutzeri could not utilize C16 individually. To gain a better understanding of the roles of the alkane nonconsumer P. stutzeri in the C16-degrading consortium, we reconstructed a two-species stoichiometric metabolic model, iBH1908, and integrated in silico prediction with the following in vitro validation, a comparative proteomics analysis, and extracellular metabolomic detection. Metabolic interactions between P. stutzeri and Dietzia sp. were successfully revealed to have importance in efficient C16 degradation. In the process, P. stutzeri survived on C16 metabolic intermediates from Dietzia sp., including hexadecanoate, 3-hydroxybutanoate, and α-ketoglutarate. In return, P. stutzeri reorganized its metabolic flux distribution to fed back acetate and glutamate to Dietzia sp. to enhance its C16 degradation efficiency by improving Dietzia cell accumulation and by regulating the expression of Dietzia succinate dehydrogenase. By using the synergistic microbial consortium of Dietzia sp. and P. stutzeri with the addition of the in silico-predicted key exchanged metabolites, diesel oil was effectively disposed of in 15 days with a removal fraction of 85.54% ± 6.42%, leaving small amounts of C15 to C20 isomers. Our finding provides a novel microbial assembling mode for efficient bioremediation or chemical production in the future.IMPORTANCE Many natural and synthetic microbial communities are composed of not only species whose biological properties are consistent with their corresponding communities but also ones whose chemophysical characteristics do not directly contribute to the performance of their communities. Even though the latter species are often essential to the microbial communities, their roles are unclear. Here, by investigation of an artificial two-member microbial consortium in n-alkane biodegradation, we showed that the microbial member without the n-alkane-degrading capability had a cross-feeding interaction with and metabolic regulation to the leading member for the synergistic n-alkane biodegradation. Our study improves the current understanding of microbial interactions. Because "assistant" microbes showed importance in communities in addition to the functional microbes, our findings also suggest a useful "assistant-microbe" principle in the design of microbial communities for either bioremediation or chemical production.

Dataset Information

Crystal Structure of TetR Family Repressor AlkX from Dietzia sp. Strain DQ12-45-1b Implicated in Biodegradation of n-Alkanes.

Publications

Crystal Structure of TetR Family Repressor AlkX from Dietzia sp. Strain DQ12-45-1b Implicated in Biodegradation of <i>n</i>-Alkanes.

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