When Coupled to Natural Transformation in<i>Acinetobacter</i>sp. Strain ADP1, PCR Mutagenesis Is Made Less Random by Mismatch Repair

Buchan, Alison; Ornston, L N

doi:10.1128/aem.71.11.7610-7612.2005

Cited by 4 publications

(3 citation statements)

References 45 publications

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“…This was first of several genes identified that encode enzymes that directly consume DMSP, the details of which are discussed below. Many of these studies were performed in cultured representatives of the well-studied roseobacters, a phylogenetically coherent clade of clade of Alphaproteobacteria that are mostly marine in origin (Buchan et al, 2005; Wagner-Dobler and Biebl, 2006). …”

Section: Introductionmentioning

confidence: 99%

Bacterial Catabolism of Dimethylsulfoniopropionate (DMSP)

2011

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Dimethylsulfoniopropionate (DMSP) is a metabolite produced primarily by marine phytoplankton and is the main precursor to the climatically important gas dimethylsulfide (DMS). DMS is released upon bacterial catabolism of DMSP, but it is not the only possible fate of DMSP sulfur. An alternative demethylation/demethiolation pathway results in the eventual release of methanethiol, a highly reactive volatile sulfur compound that contributes little to the atmospheric sulfur flux. The activity of these pathways control the natural flux of sulfur released to the atmosphere. Although these biochemical pathways and the factors that regulate them are of great interest, they are poorly understood. Only recently have some of the genes and pathways responsible for DMSP catabolism been elucidated. Thus far, six different enzymes have been identified that catalyze the cleavage of DMSP, resulting in the release of DMS. In addition, five of these enzymes appear to produce acrylate, while one produces 3-hydroxypropionate. In contrast, only one enzyme, designated DmdA, has been identified that catalyzes the demethylation reaction producing methylmercaptopropionate (MMPA). The metabolism of MMPA is performed by a series of three coenzyme-A mediated reactions catalyzed by DmdB, DmdC, and DmdD. Interestingly, Candidatus Pelagibacter ubique, a member of the SAR11 clade of Alphaproteobacteria that is highly abundant in marine surface waters, possessed functional DmdA, DmdB, and DmdC enzymes. Microbially mediated transformations of both DMS and methanethiol are also possible, although many of the biochemical and molecular genetic details are still unknown. This review will focus on the recent discoveries in the biochemical pathways that mineralize and assimilate DMSP carbon and sulfur, as well as the areas for which a comprehensive understanding is still lacking.

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Section: Introductionmentioning

confidence: 99%

Bacterial Catabolism of Dimethylsulfoniopropionate (DMSP)

2011

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“…Facile engineering of the ADP1 genome to edit and add to metabolic and sensing pathways is made possible by its constitutive expression of a competence apparatus that enables it to efficiently take up DNA from its environment in a non-sequence-specific manner (11). ADP1 can also transform fragmented and damaged DNA or mutagenized PCR products to introduce sequence variation at specific sites in its genome for optimizing engineered functions (12,13).…”

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confidence: 99%

Reduced Mutation Rate and Increased Transformability of Transposon-Free Acinetobacter baylyi ADP1-ISx

Suárez

Renda

Dasgupta

et al. 2017

Appl Environ Microbiol

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The genomes of most bacteria contain mobile DNA elements that can contribute to undesirable genetic instability in engineered cells. In particular, transposable insertion sequence (IS) elements can rapidly inactivate genes that are important for a designed function. We deleted all six copies of IS from the genome of the naturally transformable bacterium ADP1. The natural competence of ADP1 made it possible to rapidly repair deleterious point mutations that arose during strain construction. In the resulting ADP1-ISx strain, the rates of mutations inactivating a reporter gene were reduced by 7- to 21-fold. This reduction was higher than expected from the incidence of new IS insertions found during a 300-day mutation accumulation experiment with wild-type ADP1 that was used to estimate spontaneous mutation rates in the strain. The extra improvement appears to be due in part to eliminating large deletions caused by IS activity, as the point mutation rate was unchanged in ADP1-ISx. Deletion of an error-prone polymerase () and a DNA damage response regulator ( [the gene of]) from the ADP1-ISx genome did not further reduce mutation rates. Surprisingly, ADP1-ISx exhibited increased transformability. This improvement may be due to less autolysis and aggregation of the engineered cells than of the wild type. Thus, deleting IS elements from the ADP1 genome led to a greater than expected increase in evolutionary reliability and unexpectedly enhanced other key strain properties, as has been observed for other clean-genome bacterial strains. ADP1-ISx is an improved chassis for metabolic engineering and other applications. ADP1 has been proposed as a next-generation bacterial host for synthetic biology and genome engineering due to its ability to efficiently take up DNA from its environment during normal growth. We deleted transposable elements that are capable of copying themselves, inserting into other genes, and thereby inactivating them from the ADP1 genome. The resulting "clean-genome" ADP1-ISx strain exhibited larger reductions in the rates of inactivating mutations than expected from spontaneous mutation rates measured via whole-genome sequencing of lineages evolved under relaxed selection. Surprisingly, we also found that IS element activity reduces transformability and is a major cause of cell aggregation and death in wild-type ADP1 grown under normal laboratory conditions. More generally, our results demonstrate that domesticating a bacterial genome by removing mobile DNA elements that have accumulated during evolution in the wild can have unanticipated benefits.

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“…As a model soil microorganism with strong natural transformation competence, A. baylyi ADP1 has been fully sequenced (Barbe et al, 2004) and studied for decades (Hare and Gregg-Jolly, 2003;Metzgar et al, 2004;Zheng and Gregg-Jolly, 2004;Buchan and Ornston, 2005). Although A. baylyi ADP1 cells have been previously constructed as hosts for DNA damage responsive whole-cell bioreporters in environmental toxicity assessment (Ammerman and Azam, 1987;Barbe et al, 2004;Song et al, 2009;Jiang et al, 2014Jiang et al, , 2017Jia et al, 2016), the DNA damage response network in A. baylyi ADP1 is still not well-established.…”

Section: Different Behavior Of Protein Profiles In Different Carbon Smentioning

confidence: 99%

iTRAQ-Based Comparative Proteomic Analysis of Acinetobacter baylyi ADP1 Under DNA Damage in Relation to Different Carbon Sources

Jiang

Xing

et al. 2020

Front. Microbiol.

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DNA damage response allows microorganisms to repair or bypass DNA damage and maintain the genome integrity. It has attracted increasing attention but the underlying influential factors affecting DNA damage response are still unclear. In this work, isobaric tags for relative and absolute quantification (iTRAQ)-based proteomic analysis was used to investigate the influence of carbon sources on the translational response of Acinetobacter baylyi ADP1 to DNA damage. After cultivating in a nutrient-rich medium (LB) and defined media supplemented with four different carbon sources (acetate, citrate, pyruvate, and succinate), a total of 2807 proteins were identified. Among them, 84 proteins involved in stress response were significantly altered, indicating the strong influence of carbon source on the response of A. baylyi ADP1 to DNA damage and other stresses. As the first study on the comparative global proteomic changes in A. baylyi ADP1 under DNA damage across nutritional environments, our findings revealed that DNA damage response in A. baylyi ADP1 at the translational level is significantly altered by carbon source, providing an insight into the complex protein interactions across carbon sources and offering theoretical clues for further study to elucidate their general regulatory mechanism to adapt to different nutrient environments.

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When Coupled to Natural Transformation inAcinetobactersp. Strain ADP1, PCR Mutagenesis Is Made Less Random by Mismatch Repair

Cited by 4 publications

References 45 publications

Bacterial Catabolism of Dimethylsulfoniopropionate (DMSP)

Bacterial Catabolism of Dimethylsulfoniopropionate (DMSP)

Reduced Mutation Rate and Increased Transformability of Transposon-Free Acinetobacter baylyi ADP1-ISx

iTRAQ-Based Comparative Proteomic Analysis of Acinetobacter baylyi ADP1 Under DNA Damage in Relation to Different Carbon Sources

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