Effects of Zinc on Particulate Methane Monooxygenase Activity and Structure

Sirajuddin, Sarah; Barupala, Dulmini; Helling, Stefan; Marcus, Katrin; Stemmler, Timothy L.; Rosenzweig, Amy C.

doi:10.1074/jbc.m114.581363

Cited by 67 publications

(185 citation statements)

References 63 publications

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“…There are conflicting data regarding whether or not CuMbn addition affects the activity of purified pMMO (47, 54). Apo Mbn addition has been reported to increase activity (54), but surplus copper or zinc binding can inactivate pMMO (55, 56), and it may be that apo Mbn is merely removing inhibitory metals.…”

Section: Methanobactinsmentioning

confidence: 99%

Chalkophores

Kenney

Rosenzweig

2018

Annu. Rev. Biochem.

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Copper-binding metallophores, or chalkophores, play a role in microbial copper homeostasis that is analogous to that of siderophores in iron homeostasis. The best-studied chalkophores are members of the methanobactin (Mbn) family—ribosomally produced, posttranslationally modified natural products first identified as copper chelators responsible for copper uptake in methane-oxidizing bacteria. To date, Mbns have been characterized exclusively in those species, but there is genomic evidence for their production in a much wider range of bacteria. This review addresses the current state of knowledge regarding the function, biosynthesis, transport, and regulation of Mbns. While the roles of several proteins in these processes are supported by substantial genetic and biochemical evidence, key aspects of Mbn manufacture, handling, and regulation remain unclear. In addition, other natural products that have been proposed to mediate copper uptake as well as metallophores that have biologically relevant roles involving copper binding, but not copper uptake, are discussed.

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Section: Methanobactinsmentioning

confidence: 99%

Chalkophores

Kenney

Rosenzweig

2018

Annu. Rev. Biochem.

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“…50,96 Moreover, zinc inhibits pMMO, likely by binding in the crystallographic site, leading to the hypothesis that the coordinating residues, which are conserved, could be involved in proton transfer. 97 It is unclear if copper binding at this site is functionally important. Samples of pMMOs in which copper is observed in the PmoC site have been prepared in the presence of extra copper, while pMMOs that have not been treated with additional copper require zinc to crystallize.…”

Section: Aerobic Methane Oxidationmentioning

confidence: 99%

Methane-Oxidizing Enzymes: An Upstream Problem in Biological Gas-to-Liquids Conversion

2016

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Biological conversion of natural gas to liquids (Bio-GTL) represents an immense economic opportunity. In nature, aerobic methanotrophic bacteria and anaerobic archaea are able to selectively oxidize methane using methane monooxygenase (MMO) and methyl coenzyme M reductase (MCR) enzymes. Although significant progress has been made toward genetically manipulating these organisms for biotechnological applications, the enzymes themselves are slow, complex, and not recombinantly tractable in traditional industrial hosts. With turnover numbers of 0.16–13 s−1, these enzymes pose a considerable upstream problem in the biological production of fuels or chemicals from methane. Methane oxidation enzymes will need to be engineered to be faster to enable high volumetric productivities; however, efforts to do so and to engineer simpler enzymes have been minimally successful. Moreover, known methane-oxidizing enzymes have different expression levels, carbon and energy efficiencies, require auxiliary systems for biosynthesis and function, and vary considerably in terms of complexity and reductant requirements. The pros and cons of using each methane-oxidizing enzyme for Bio-GTL are considered in detail. The future for these enzymes is bright, but a renewed focus on studying them will be critical to the successful development of biological processes that utilize methane as a feedstock.

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“…Three additional crystal structures of pMMO have been published, showing only asingle Cu ion in site A. [10] Unfortunately,t he crystal structures did not settle the location and composition of the pMMO active site.Computational studies have supported site Aa nd have indicated that both mono-and dinuclear Cu sites are reactive enough to oxidise CH 4 . [3,11] On the other hand, Chan and co-workers have argued that the active site instead consists of three Cu ions,not observed in the crystal structure,but coordinated by residues from PmoA and PmoC.…”

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Quantum Refinement Does Not Support Dinuclear Copper Sites in Crystal Structures of Particulate Methane Monooxygenase

et al. 2017

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Particulate methane monooxygenase (pMMO) is one of the few enzymes that can activate methane.T he metal content of this enzyme has been highly controversial, with suggestions of ad inuclear Fe site or mono-, di-, or trinuclear Cu sites.C rystal structures have shown am ono-or dinuclear Cu site,b ut the resolution was low and the geometry of the dinuclear site unusual. We have employed quantum refinement (crystallographic refinement enhanced with quantum-mechanical calculations) to improve the structure of the active site.W e compared an umber of different mono-and dinuclear geometries,i ns ome cases enhanced with more protein ligands or one or two water molecules,t od etermine which structure fits two sets of crystallographic raw data best. In all cases,the best results were obtained with mononuclear Cu sites,occasionally with an extra water molecule.Thus,weconclude that there is no crystallographic support for adinuclear Cu site in pMMO.Methane is one of the most difficult organic substrates to activate,w ith aC À Hb ond dissociation energy of 435 kJ mol À1 .[1] In biology,t here are only two groups of enzymes that can perform this reaction. Thef irst one is composed of the soluble methane monooxygenases, [2] which contain ad inuclear Fe active site,s imilar to that of the ribonucleotide reductases.T his is aw ell-characterised group of enzymes,a nd their reaction mechanism is understood in some detail.[2] Thesecond group contains the particulate (i.e., membrane-bound) methane monooxygenases (pMMO), which are the predominant enzymes in methanotrophic bacteria, except under copper-limiting conditions. [1,3] The pMMOs have been more difficult to characterise,a nd the metal content has been highly controversial, with suggestions of dinuclear Fe centres as well as mono-, di-, and trinuclear Cu sites. [1,[4][5][6] In 2005, the first crystal structure of ap MMO was published (2.8 resolution), showing aheterotrimeric a 3 b 3 g 3 structure with ad inuclear and am ononuclear Cu site in the soluble periplasmic parts of the PmoB subunit and am onomeric Zn site in the PmoC subunit within the membrane (these three metal sites will be called A, B, and Ci nt he following;F igure 1). [7] Site Awas suggested to be the active site and involves three histidine residues,which are conserved in all pMMO sequences,except for those from one phylum of bacteria.[1] All three residues interact with the metals via their side-chain imidazole group,but one of them is the first residue of PmoB and the amino-terminal Natom also coordinates to one of the metals.T his gives ab inding site similar to the histidine brace in the lytic polysaccharide monooxygenase enzymes, [8] although that site involves only two His ligands Figure 1. a) Trimeric structure of pMMO (3RGB) [9] with metal sites ACindicated. Cu ions in yellow,Z nions in red;PmoA, PmoB, and PmoC subunits in magenta, green, and cyan, respectively.Geometry of Cu site Ai nprotomers 1(b) and 2( c), involving three histidine residues and the amino-terminal group. Cu ions are shown ...

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Effects of Zinc on Particulate Methane Monooxygenase Activity and Structure

Cited by 67 publications

References 63 publications

Chalkophores

Chalkophores

Methane-Oxidizing Enzymes: An Upstream Problem in Biological Gas-to-Liquids Conversion

Quantum Refinement Does Not Support Dinuclear Copper Sites in Crystal Structures of Particulate Methane Monooxygenase

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