Autoinhibition and Signaling by the Switch II Motif in the G-protein Chaperone of a Radical B12 Enzyme

Lofgren, Michael; Koutmos, Markos; Banerjee, Ruma

doi:10.1074/jbc.m113.499970

Cited by 18 publications

(26 citation statements)

References 49 publications

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“…The latter synthesizes AdoCbl and delivers it to its target mutase (11). In this study, we demonstrate its additional role in cofactor offloading from an inactivated mutase and provide molecular insights into a more complex pattern of allosteric regulation of AdoCbl loading and repair than was previously known (8,11,13,16,17).…”

Section: Discussionmentioning

confidence: 75%

“…Two proteins, adenosyltransferase (ATR) (9) and a G-protein chaperone (10), are needed for cofactor repair and reloading. The Methylobacterium extorquens orthologs of MCM, ATR and the G-protein (known as MeaB), have been characterized extensively (8,(11)(12)(13)(14)(15)(16)(17), and our understanding of the mechanism of cofactor repair derives primarily from studies on this system. The importance of the chaperonedependent cofactor loading and repair mechanisms is underscored by the existence of disease-causing mutations in both human ATR and the G-protein chaperone (18,19).…”

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confidence: 99%

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Cofactor Editing by the G-protein Metallochaperone Domain Regulates the Radical B12 Enzyme IcmF

Zhu

Kitanishi

Twahir

et al. 2017

Journal of Biological Chemistry

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Edited by George N. DeMartinoIcmF is a 5-deoxyadenosylcobalamin (AdoCbl)-dependent enzyme that catalyzes the carbon skeleton rearrangement of isobutyryl-CoA to butyryl-CoA. It is a bifunctional protein resulting from the fusion of a G-protein chaperone with GTPase activity and the cofactor-and substrate-binding mutase domains with isomerase activity. IcmF is prone to inactivation during catalytic turnover, thus setting up its dependence on a cofactor repair system. Herein, we demonstrate that the GTPase activity of IcmF powers the ejection of the inactive cob(II)alamin cofactor and requires the presence of an acceptor protein, adenosyltransferase, for receiving it. Adenosyltransferase in turn converts cob(II)alamin to AdoCbl in the presence of ATP and a reductant. The repaired cofactor is then reloaded onto IcmF in a GTPase-gated step. The mechanistic details of cofactor loading and offloading from the AdoCbl-dependent IcmF are distinct from those of the better characterized and homologous methylmalonyl-CoA mutase/G-protein chaperone system. Acyl-CoA mutases are a group of enzymes that utilize 5Ј-deoxyadenosylcobalamin (AdoCbl 4 or coenzyme B 12 ) as a cofactor for catalyzing carbon skeleton rearrangement reactions (1, 2). Isobutyryl-CoA mutase is a member of this enzyme family that is found in bacteria and catalyzes the reversible rearrangement of isobutyryl-and butyryl-CoA mutase ( Fig. 1A) (3). A chemically similar interconversion of methylmalonyl-CoA and succinyl-CoA is catalyzed by methylmalonyl-CoA mutase (MCM) (4), which is the only AdoCbl-dependent enzyme found in mammals. AdoCbl-dependent isomerization reactions are initiated by the homolytic cleavage of the cobalt-carbon bond of AdoCbl yielding cob(II)alamin and the working 5Ј-deoxyadenosyl radical, which initiates the chemical reaction by hydrogen atom abstraction from the substrate. AdoCbl thus serves as a latent radical reservoir (5), and substrate binding accelerates the homolytic cleavage rate in AdoCbl-dependent enzymes by a factor of ϳ10 12 (6, 7). At the end of each catalytic cycle, the 5Ј-deoxyadenosyl radical and cob(II)alamin recombine, thus regenerating the resting AdoCbl form of the cofactor (Fig. 1A). During catalytic turnover, the occasional loss of the 5Ј-deoxyadenosine moiety renders MCM prone to inactivation (8) and necessitates its dependence on a repair system for reentry into the catalytic cycle. Two proteins, adenosyltransferase (ATR) (9) and a G-protein chaperone (10), are needed for cofactor repair and reloading. The Methylobacterium extorquens orthologs of MCM, ATR and the G-protein (known as MeaB), have been characterized extensively (8,(11)(12)(13)(14)(15)(16)(17), and our understanding of the mechanism of cofactor repair derives primarily from studies on this system. The importance of the chaperonedependent cofactor loading and repair mechanisms is underscored by the existence of disease-causing mutations in both human ATR and the G-protein chaperone (18,19).IcmF is a naturally occurring fusion in which the G-protein chapero...

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Section: Discussionmentioning

confidence: 75%

mentioning

confidence: 99%

Cofactor Editing by the G-protein Metallochaperone Domain Regulates the Radical B12 Enzyme IcmF

Zhu

Kitanishi

Twahir

et al. 2017

Journal of Biological Chemistry

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“…2A). Asp249 is in proximity to the Mg 2+ -binding site and seems ideally positioned to activate a water molecule for GTP hydrolysis, as predicted previously based on homology to HypB (32). Arg265, which was disordered in structures of MeaB alone, interacts with the phosphate groups of the bound GDP and could help stabilize the negative charge that builds up during GTP hydrolysis, thus playing the role of a cis-acting Arg finger.…”

Section: Resultsmentioning

confidence: 89%

Visualization of a radical B ₁₂ enzyme with its G-protein chaperone

Jost¹,

Cracan

Hubbard³

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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G-protein metallochaperones ensure fidelity during cofactor assembly for a variety of metalloproteins, including adenosylcobalamin (AdoCbl)-dependent methylmalonyl-CoA mutase and hydrogenase, and thus have both medical and biofuel development applications. Here, we present crystal structures of IcmF, a natural fusion protein of AdoCbl-dependent isobutyryl-CoA mutase and its corresponding G-protein chaperone, which reveal the molecular architecture of a G-protein metallochaperone in complex with its target protein.These structures show that conserved G-protein elements become ordered upon target protein association, creating the molecular pathways that both sense and report on the cofactor loading state. Structures determined of both apo-and holo-forms of IcmF depict both open and closed enzyme states, in which the cofactor-binding domain is alternatively positioned for cofactor loading and for catalysis. Notably, the G protein moves as a unit with the cofactorbinding domain, providing a visualization of how a chaperone assists in the sequestering of a precious cofactor inside an enzyme active site.

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“…G3E family GTPases, like most G-proteins, bind GDP and GTP using several short, conserved loops that often occur between an α-helix and a β-strand and often at a dimer interface, numbered G1–G5 [28] (Figure 2). G1, also known as the P-loop or Walker A motif binds to the phosphates in GDP.…”

Section: Metallochaperonesmentioning

confidence: 99%

Metallochaperones and metalloregulation in bacteria

Capdevila

Edmonds

Giedroc

2017

Essays in Biochemistry

110

117

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Bacterial transition metal homoeostasis or simply ‘metallostasis’ describes the process by which cells control the intracellular availability of functionally required metal cofactors, from manganese (Mn) to zinc (Zn), avoiding bothmetal deprivation and toxicity. Metallostasis is an emerging aspect of the vertebrate host–pathogen interface that is defined by a ‘tug-of-war’ for biologically essential metals and provides the motivation for much recent work in this area. The host employs a number of strategies to starve the microbial pathogen of essential metals, while for others attempts to limit bacterial infections by leveraging highly competitive metals. Bacteria must be capable of adapting to these efforts to remodel the transition metal landscape and employ highly specialized metal sensing transcriptional regulators, termed metalloregulatory proteins, and metallochaperones, that allocate metals to specific destinations, to mediate this adaptive response. In this essay, we discuss recent progress in our understanding of the structural mechanisms and metal specificity of this adaptive response, focusing on energy-requiring metallochaperones that play roles in the metallocofactor active site assembly in metalloenzymes and metallosensors, which govern the systems-level response to metal limitation and intoxication.

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Autoinhibition and Signaling by the Switch II Motif in the G-protein Chaperone of a Radical B12 Enzyme

Cited by 18 publications

References 49 publications

Cofactor Editing by the G-protein Metallochaperone Domain Regulates the Radical B12 Enzyme IcmF

Cofactor Editing by the G-protein Metallochaperone Domain Regulates the Radical B12 Enzyme IcmF

Visualization of a radical B ₁₂ enzyme with its G-protein chaperone

Metallochaperones and metalloregulation in bacteria

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Autoinhibition and Signaling by the Switch II Motif in the G-protein Chaperone of a Radical B12 Enzyme

Cited by 18 publications

References 49 publications

Cofactor Editing by the G-protein Metallochaperone Domain Regulates the Radical B12 Enzyme IcmF

Cofactor Editing by the G-protein Metallochaperone Domain Regulates the Radical B12 Enzyme IcmF

Visualization of a radical B 12 enzyme with its G-protein chaperone

Metallochaperones and metalloregulation in bacteria

Contact Info

Product

Resources

About

Visualization of a radical B ₁₂ enzyme with its G-protein chaperone