Competition between Metals for Binding to Methanobactin Enables Expression of Soluble Methane Monooxygenase in the Presence of Copper

Kalidass, Bhagyalakshmi; Haque, Muhammad Farhan Ul; Baral, Bipin S.; DiSpirito, Alan A.; Semrau, Jeremy D.

doi:10.1128/aem.03151-14

Cited by 27 publications

(24 citation statements)

References 41 publications

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“…As found previously (35,36,39), the addition of copper reduced mmoX expression by ϳ4 orders of magnitude ( Fig. 2A), while pmoA expression increased ϳ54-fold (Fig.…”

Section: Resultssupporting

confidence: 63%

“…S2 in the supplemental material). As found earlier (36,39), copper associated with biomass significantly increased (approximately three orders of magnitude; P Ͻ 6 ϫ 10 Ϫ3 ) with the addition of copper (Fig. 1A).…”

Section: Resultssupporting

confidence: 57%

“…Total RNA extraction from M. trichosporium OB3b was performed as described earlier (36). Briefly, 2.5 ml of stop solution (5% buffer equilibrated phenol [pH 7.3] in ethanol) was first added to cultures (22.5 ml) to stop synthesis of new mRNA.…”

Section: Methodsmentioning

confidence: 99%

“…Copper and cerium associated with biomass and remaining in the supernatant after growth were determined as described earlier (36). Briefly, cultures of M. trichosporium OB3b were centrifuged at 5,000 ϫ g for 10 min at 4°C.…”

Section: Methodsmentioning

confidence: 99%

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Cerium Regulates Expression of Alternative Methanol Dehydrogenases in Methylosinus trichosporium OB3b

Haque

Kalidass

Bandow

et al. 2015

Appl Environ Microbiol

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bMethanotrophs have multiple methane monooxygenases that are well known to be regulated by copper, i.e., a "copper switch." At low copper/biomass ratios the soluble methane monooxygenase (sMMO) is expressed while expression and activity of the particulate methane monooxygenase (pMMO) increases with increasing availability of copper. In many methanotrophs there are also multiple methanol dehydrogenases (MeDHs), one based on Mxa and another based on Xox. Mxa-MeDH is known to have calcium in its active site, while Xox-MeDHs have been shown to have rare earth elements in their active site. We show here that the expression levels of Mxa-MeDH and Xox-MeDH in Methylosinus trichosporium OB3b significantly decreased and increased, respectively, when grown in the presence of cerium but the absence of copper compared to the absence of both metals. Expression of sMMO and pMMO was not affected. In the presence of copper, the effect of cerium on gene expression was less significant, i.e., expression of Mxa-MeDH in the presence of copper and cerium was slightly lower than in the presence of copper alone, but Xox-MeDH was again found to increase significantly. As expected, the addition of copper caused sMMO and pMMO expression levels to significantly decrease and increase, respectively, but the simultaneous addition of cerium had no discernible effect on MMO expression. As a result, it appears Mxa-MeDH can be uncoupled from methane oxidation by sMMO in M. trichosporium OB3b but not from pMMO. It is well known that microorganisms have diverse mechanisms to sense and respond to metals in their environment. These mechanisms typically include strategies to regulate gene expression in response to the presence or absence of metals such as copper, zinc, iron, manganese, arsenic, and mercury (see, for example, references 1, 2, 3 and 4). One such phenomenon is the "copper switch" in methanotrophs. That is, these microbes utilize methane as their sole growth substrate but have two different monooxygenases for the initial oxidation of methane to methanol. One, the particulate methane monooxygenase (pMMO), is found in the intracytoplasmic membranes of these microbes, and its expression and activity increases with increasing availability of copper. The second, the soluble methane monooxygenase (sMMO), is found in the cytoplasm and is only expressed when copper is unavailable (5). These two forms of MMO have very different structures, activities, and substrate ranges (5-14), and so careful consideration of the form of MMO expressed is critical for understanding methanotrophic ecology, as well for various applications of methanotrophy, including the removal of chlorinated solvents and methane, a potent greenhouse gas (5, 10, 12, 15).Further, interest in the commercial application of methanotrophy has dramatically accelerated in recent years, in part due to increased methane supplies given advances in hydraulic fracturing of shale formations. As a result, methane prices have become quite low, with the wellhead price of natural gas dropping fro...

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“…As found previously (35,36,39), the addition of copper reduced mmoX expression by ϳ4 orders of magnitude ( Fig. 2A), while pmoA expression increased ϳ54-fold (Fig.…”

Section: Resultssupporting

confidence: 63%

Section: Resultssupporting

confidence: 57%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Cerium Regulates Expression of Alternative Methanol Dehydrogenases in Methylosinus trichosporium OB3b

Haque

Kalidass

Bandow

et al. 2015

Appl Environ Microbiol

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“…Methylosinus trichosporium OB3b was grown in nitrate minimal salts (NMS) medium (32) at 30°C without any added copper or with 1 M copper as CuCl 2 . The background copper concentration in standard NMS medium was 0.03 Ϯ 0.01 M, as determined using atomic absorption spectroscopy (33). Methanobactin from Methylocystis sp.…”

Section: Methodsmentioning

confidence: 99%

Methanobactin from Methylocystis sp. Strain SB2 Affects Gene Expression and Methane Monooxygenase Activity in Methylosinus trichosporium OB3b

Haque

Kalidass

Vorobev

et al. 2015

Appl Environ Microbiol

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bMethanotrophs can express a cytoplasmic (soluble) methane monooxygenase (sMMO) or membrane-bound (particulate) methane monooxygenase (pMMO). Expression of these MMOs is strongly regulated by the availability of copper. Many methanotrophs have been found to synthesize a novel compound, methanobactin (Mb), that is responsible for the uptake of copper, and methanobactin produced by Methylosinus trichosporium OB3b plays a key role in controlling expression of MMO genes in this strain. As all known forms of methanobactin are structurally similar, it was hypothesized that methanobactin from one methanotroph may alter gene expression in another. When Methylosinus trichosporium OB3b was grown in the presence of 1 M CuCl 2 , expression of mmoX, encoding a subunit of the hydroxylase component of sMMO, was very low. mmoX expression increased, however, when methanobactin from Methylocystis sp. strain SB2 (SB2-Mb) was added, as did whole-cell sMMO activity, but there was no significant change in the amount of copper associated with M. trichosporium OB3b. If M. trichosporium OB3b was grown in the absence of CuCl 2 , the mmoX expression level was high but decreased by several orders of magnitude if copper prebound to SB2-Mb (Cu-SB2-Mb) was added, and biomass-associated copper was increased. Exposure of Methylosinus trichosporium OB3b to SB2-Mb had no effect on expression of mbnA, encoding the polypeptide precursor of methanobactin in either the presence or absence of CuCl 2 . mbnA expression, however, was reduced when Cu-SB2-Mb was added in both the absence and presence of CuCl 2 . These data suggest that methanobactin acts as a general signaling molecule in methanotrophs and that methanobactin "piracy" may be commonplace. Methanotrophs are distinguished from other microorganisms by their ability to utilize methane as a sole carbon and energy source yet are phylogenetically and physiologically diverse. Microbial methane oxidation can be coupled to a variety of terminal electron acceptors, including oxygen, sulfate, nitrate, and nitrite (1-4). The aerobic methanotrophs are typically mesophilic and group phylogenetically within the Gammaproteobacteria and Alphaproteobacteria (1). Thermo-and meso-acidophilic aerobic methanotrophs, however, that grow at pH Ͻ3 and at optimal temperatures ranging from 35°C to greater than 50°C have also been discovered in the phylum Verrucomicrobia (5-9). Further, novel oxygenic methanotrophs that couple methane oxidation to nitrite reduction have been reported, e.g., "Candidatus Methylomirabilis oxyfera" that generates oxygen from a unique denitrification pathway, which is then used for methane oxidation (2). Aerobic methanotrophs are found in many environments, e.g., freshwater and marine sediments, bogs, forest, and agricultural soils, among other locations (1, 2, 5-11).These microorganisms have been extensively studied for many different reasons, including the fact that they play a key role in the global carbon cycle. All aerobic methanotrophs employ the enzyme methane monooxygenase (MMO) to co...

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Effects of copper on expression of methane monooxygenases, trichloroethylene degradation, and community structure in methanotrophic consortia

Xing

Zhao

Zhang

et al. 2018

Engineering in Life Sciences

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Copper plays a key role in regulating the expression of enzymes that promote biodegradation of contaminants in methanotrophic consortia (MC). Here, we utilized MC isolated from landfill cover to investigate cometabolic degradation of trichloroethylene (TCE) at nine different copper (Cu2+) concentrations. The results demonstrated that an increase in Cu2+ concentration from 0 to 15 μM altered the specific first‐order rate constant k1,TCE, the expression levels of methane monooxygenase (pmoA and mmoX) genes, and the specific activity of soluble methane monooxygenase (sMMO). High efficiency TCE degradation (95%) and the expression levels of methane monooxygenase (MMO) were detected at a Cu2+ concentration of 0.03 μM. Notably, sMMO‐specific activity ranged from 74.41 nmol/(mgcell h) in 15 μM Cu2+ to 654.99 nmol/(mgcell h) in 0.03 μM Cu2+, which contrasts with cultures of pure methanotrophs in which sMMO activity is depressed at high Cu2+ concentrations, indicating a special regulatory role for Cu2+ in MC. The results of MiSeq pyrosequencing indicated that higher Cu2+ concentrations stimulated the growth of methanotrophic microorganisms in MC. These findings have important implications for the elucidation of copper‐mediated regulatory mechanisms in MC.

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Competition between Metals for Binding to Methanobactin Enables Expression of Soluble Methane Monooxygenase in the Presence of Copper

Cited by 27 publications

References 41 publications

Cerium Regulates Expression of Alternative Methanol Dehydrogenases in Methylosinus trichosporium OB3b

Cerium Regulates Expression of Alternative Methanol Dehydrogenases in Methylosinus trichosporium OB3b

Methanobactin from Methylocystis sp. Strain SB2 Affects Gene Expression and Methane Monooxygenase Activity in Methylosinus trichosporium OB3b

Effects of copper on expression of methane monooxygenases, trichloroethylene degradation, and community structure in methanotrophic consortia

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