scholarly journals Generation of a Gluconobacter oxydans knockout collection for improved extraction of rare earth elements

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Alexa M. Schmitz ◽  
Brooke Pian ◽  
Sean Medin ◽  
Matthew C. Reid ◽  
Mingming Wu ◽  
...  
Keyword(s):  
Rare Earth ◽  
Rare Earth Elements ◽  
Transport System ◽  
Glucose Dehydrogenase ◽  
Single Gene ◽  
Gluconobacter Oxydans ◽  
Pyrroloquinoline Quinone ◽  
Whole Genome ◽  
Membrane Bound ◽  
Specific Transport System

AbstractBioleaching of rare earth elements (REEs), using microorganisms such as Gluconobacter oxydans, offers a sustainable alternative to environmentally harmful thermochemical extraction, but is currently not very efficient. Here, we generate a whole-genome knockout collection of single-gene transposon disruption mutants for G. oxydans B58, to identify genes affecting the efficacy of REE bioleaching. We find 304 genes whose disruption alters the production of acidic biolixiviant. Disruption of genes underlying synthesis of the cofactor pyrroloquinoline quinone (PQQ) and the PQQ-dependent membrane-bound glucose dehydrogenase nearly eliminates bioleaching. Disruption of phosphate-specific transport system genes enhances bioleaching by up to 18%. Our results provide a comprehensive roadmap for engineering the genome of G. oxydans to further increase its bioleaching efficiency.

scholarly journals Cloning and Nucleotide Sequencing of the Membrane-Bound l-Sorbosone Dehydrogenase Gene of Acetobacter liquefaciens IFO 12258 and Its Expression in Gluconobacter oxydans

1995 ◽  
Vol 61 (5) ◽  
pp. 2069-2069
Author(s):  
M Shinjoh ◽  
N Tomiyama ◽  
A Asakura ◽  
T Hoshino
Keyword(s):  
Alcohol Dehydrogenase ◽  
Glucose Dehydrogenase ◽  
Gluconobacter Oxydans ◽  
Pyrroloquinoline Quinone ◽  
Nucleotide Sequencing ◽  
Common Region ◽  
Quinone Binding ◽  
Dehydrogenase Gene ◽  
Membrane Bound

Volume 61, no. 2, p. 419, column 1, lines 15-19: this sentence should read as follows. "The alcohol dehydrogenase and glucose dehydrogenase have a common region reported to be related to pyrroloquinoline quinone binding (2, 10), but SNDH does not contain such a region, indicating that SNDH is not a quinoprotein." Page 419, column 2, line 12: "(Table 4)" should read "(Table 3)." [This corrects the article on p. 413 in vol. 61.].

scholarly journals Gluconobacter oxydans Knockout Collection Finds Improved Rare Earth Element Extraction

2021 ◽  
Author(s):  
Alexa M. Schmitz ◽  
Brooke Pian ◽  
Sean Medin ◽  
Matthew C. Reid ◽  
Mingming Wu ◽  
...  
Keyword(s):  
Rare Earth ◽  
Acid Production ◽  
Glucose Dehydrogenase ◽  
Gluconobacter Oxydans ◽  
Extraction Methods ◽  
Promising Alternative ◽  
Energy Technologies ◽  
Thermochemical Processes ◽  
Membrane Bound ◽  
Critical Components

Rare earth elements (REE) are critical components of our technological society and essential for renewable energy technologies. Traditional thermochemical processes to extract REE from mineral ores or recycled materials are costly and environmentally harmful, and thus more sustainable extraction methods require exploration. Bioleaching offers a promising alternative to conventional REE extraction, and is already used to extract 5% of the world's gold, and approximately 15% of the world's copper supply. However, the performance of REE bioleaching lags far behind thermochemical processes. Despite this, to the best of our knowledge no genetic engineering strategies have yet been used to enhance REE bioleaching, and little is known of the genetics that confer this capability. Here we build a whole genome knockout collection for Gluconobacter oxydans B58, one of the most promising organisms for REE bioleaching, and use it to comprehensively characterize the genomics of REE bioleaching. In total, we find 304 genes that notably alter production of G. oxydans' acidic biolixiviant, including 165 that hold up under statistical comparison with wild-type. The two most impactful groups of genes involved in REE bioleaching have opposing influences on acid production and REE bioleaching. Disruption of genes underlying synthesis of the cofactor pyrroloquinoline quinone (PQQ) and the PQQ-dependent membrane-bound glucose dehydrogenase all but eliminates bioleaching. In contrast, disruption of the phosphate-specific transport system accelerates acid production and enhances bioleaching. We identified 6 disruption mutants, that increase bioleaching by at least 11%. Most significantly, disruption of pstC, encoding part of the phosphate -specific transporter, pstSCAB, enhances bioleaching by 18%. Taken together, these results give a comprehensive roadmap for engineering multiple sites in the genome of G. oxydans to further increase its bioleaching efficiency.

scholarly journals Knockout and Overexpression of Pyrroloquinoline Quinone Biosynthetic Genes in Gluconobacter oxydans 621H

2006 ◽  
Vol 188 (21) ◽  
pp. 7668-7676 ◽  
Author(s):  
Tina Hölscher ◽  
Helmut Görisch
Keyword(s):  
Energy Source ◽  
Wild Type Strain ◽  
Gluconobacter Oxydans ◽  
Analysis Data ◽  
Pyrroloquinoline Quinone ◽  
Vital Role ◽  
Reverse Transcription Pcr ◽  
Membrane Bound ◽  
Pqq Biosynthesis

ABSTRACT In Gluconobacter oxydans, pyrroloquinoline quinone (PQQ) serves as the cofactor for various membrane-bound dehydrogenases that oxidize sugars and alcohols in the periplasm. Proteins for the biosynthesis of PQQ are encoded by the pqqABCDE gene cluster. Our reverse transcription-PCR and promoter analysis data indicated that the pqqA promoter represents the only promoter within the pqqABCDE cluster of G. oxydans 621H. PQQ overproduction in G. oxydans was achieved by transformation with the plasmid-carried pqqA gene or the complete pqqABCDE cluster. A G. oxydans mutant unable to produce PQQ was obtained by site-directed disruption of the pqqA gene. In contrast to the wild-type strain, the pqqA mutant did not grow with d-mannitol, d-glucose, or glycerol as the sole energy source, showing that in G. oxydans 621H, PQQ is essential for growth with these substrates. Growth of the pqqA mutant, however, was found with d-gluconate as the energy source. The growth behavior of the pqqA mutant correlated with the presence or absence of the respective PQQ-dependent membrane-bound dehydrogenase activities, demonstrating the vital role of these enzymes in G. oxydans metabolism. A different PQQ-deficient mutant was generated by Tn5 transposon mutagenesis. This mutant showed a defect in a gene with high homology to the Escherichia coli tldD gene, which encodes a peptidase. Our results indicate that the tldD gene in G. oxydans 621H is involved in PQQ biosynthesis, possibly with a similar function to that of the pqqF genes found in other PQQ-synthesizing bacteria.

scholarly journals A tunable l-arabinose-inducible expression plasmid for the acetic acid bacterium Gluconobacter oxydans

2020 ◽  
Vol 104 (21) ◽  
pp. 9267-9282
Author(s):  
Philipp Moritz Fricke ◽  
Tobias Link ◽  
Jochem Gätgens ◽  
Christiane Sonntag ◽  
Maike Otto ◽  
...  
Keyword(s):  
Acetic Acid ◽  
Stationary Phase ◽  
Gene Knockout ◽  
Glucose Dehydrogenase ◽  
Gluconobacter Oxydans ◽  
Acetic Acid Bacteria ◽  
Acetic Acid Bacterium ◽  
Basal Expression ◽  
Fold Induction ◽  
Membrane Bound

Abstract The acetic acid bacterium (AAB) Gluconobacter oxydans incompletely oxidizes a wide variety of carbohydrates and is therefore used industrially for oxidative biotransformations. For G. oxydans, no system was available that allows regulatable plasmid-based expression. We found that the l-arabinose-inducible PBAD promoter and the transcriptional regulator AraC from Escherichia coli MC4100 performed very well in G. oxydans. The respective pBBR1-based plasmids showed very low basal expression of the reporters β-glucuronidase and mNeonGreen, up to 480-fold induction with 1% l-arabinose, and tunability from 0.1 to 1% l-arabinose. In G. oxydans 621H, l-arabinose was oxidized by the membrane-bound glucose dehydrogenase, which is absent in the multi-deletion strain BP.6. Nevertheless, AraC-PBAD performed similar in both strains in the exponential phase, indicating that a gene knockout is not required for application of AraC-PBAD in wild-type G. oxydans strains. However, the oxidation product arabinonic acid strongly contributed to the acidification of the growth medium in 621H cultures during the stationary phase, which resulted in drastically decreased reporter activities in 621H (pH 3.3) but not in BP.6 cultures (pH 4.4). These activities could be strongly increased quickly solely by incubating stationary cells in d-mannitol-free medium adjusted to pH 6, indicating that the reporters were hardly degraded yet rather became inactive. In a pH-controlled bioreactor, these reporter activities remained high in the stationary phase (pH 6). Finally, we created a multiple cloning vector with araC-PBAD based on pBBR1MCS-5. Together, we demonstrated superior functionality and good tunability of an AraC-PBAD system in G. oxydans that could possibly also be used in other AAB. Key points • We found the AraC-PBADsystem from E. coli MC4100 was well tunable in G. oxydans. •  In the absence of AraC orl-arabinose, expression from PBADwas extremely low. • This araC-PBADsystem could also be fully functional in other acetic acid bacteria.

scholarly journals Membrane-Bound Pyrroloquinoline Quinone-Dependent Dehydrogenase in Gluconobacter oxydans M5, Responsible for Production of 6-(2-Hydroxyethyl) Amino-6-Deoxy-l-Sorbose

2008 ◽  
Vol 74 (16) ◽  
pp. 5250-5253 ◽  
Author(s):  
Xue-Peng Yang ◽  
Liu-Jing Wei ◽  
Jin-Ping Lin ◽  
Bo Yin ◽  
Dong-Zhi Wei
Keyword(s):  
Gene Disruption ◽  
Gluconobacter Oxydans ◽  
Pyrroloquinoline Quinone ◽  
Sorbitol Dehydrogenase ◽  
Antidiabetic Drug ◽  
Dehydrogenase Gene ◽  
Membrane Bound

ABSTRACT A membrane-bound protein purified from Gluconobacter oxydans M5 was confirmed to be a pyrroloquinoline quinone-dependent d-sorbitol dehydrogenase. Gene disruption and complementation experiments demonstrated that this enzyme is responsible for the oxidation of 1-(2-hydroxyethyl) amino-1-deoxy-d-sorbitol (1NSL) to 6-(2-hydroxyethyl) amino-6-deoxy-l-sorbose (6NSE), which is the precursor of an antidiabetic drug, miglitol.

2-Deoxyglucose Transportation Via Passive Diffusion and its Oxidation, Not Phosphorylation to 2-Deoxygluconic Acid by Pseudomonas aeruginosa

1971 ◽  
Vol 49 (5) ◽  
pp. 606-613 ◽  
Author(s):  
R. G. Eagon
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Active Transport ◽  
Transport System ◽  
Glucose Dehydrogenase ◽  
Passive Diffusion ◽  
Suggestive Evidence ◽  
System 2 ◽  
Membrane Bound ◽  
Active Transport System ◽  
Oxidation Of Glucose

The transport and catabolism of 2-deoxyglucose by Pseudomonas aeruginosa were studied. 2-Deoxyglucose was taken up and oxidized by glucose-grown cells at a rate approaching that of the uptake and oxidation of glucose. However, 2-deoxyglucose entered these cells via passive diffusion while glucose entered via an inducible active transport system. 2-Deoxyglucose was oxidized stoichiometrically, presumably by glucose dehydrogenase, to 2-deoxygluconic acid which, in turn, diffused out of the cells. No phosphorylated intermediates were detected. Glucose dehydrogenase was induced by cultivating the cells on glucose. There was suggestive evidence that the membrane-bound glucose dehydrogenase provided energy for the transport of glucose.

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