Pathogenic Mutations Differentially Affect the Catalytic Activities of the Human B12-processing Chaperone CblC and Increase Futile Redox Cycling

Gherasim, Carmen; Ruetz, Markus; Li, Zhu; Hudolin, Stephanie; Banerjee, Ruma

doi:10.1074/jbc.m115.637132

Cited by 32 publications

(39 citation statements)

References 51 publications

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“…Another observation in this study is both of these pathogenic variants do not significantly affect the decyanation activity, and the authors suggested that individuals with these pathogenic variants might respond to cyanocobalamin. 28 The mild clinical and biochemical phenotype associated with the c.482G>A (p.Arg161Gln) pathogenic variant could be explained by findings in these two studies, including intermediate stability of the mutant protein and the stabilization by an increase in cobalamin concentration. Moreover, Learner-Ellis et al showed higher expression of the c.482G>A(p.Arg161Gln) mutant protein compared to the c.271dupA (p.Arg91LysfsX14) pathogenic variant 12 .…”

Section: Discussionmentioning

confidence: 91%

“…This residue is located in a cavity near the corrin ring and does not make direct contact with cobalamin 28,29 . It has been proposed that this arginine residue is important for binding of glutathione 29 , with glutathione serving as an electron donor in the dealkylation reaction of adenosylcobalamin and methylcobalamin.…”

Section: Discussionmentioning

confidence: 99%

“…These results suggested that the c.482G>A (p.Arg161Gln) pathogenic variant causes decreased stability of the mutant protein and less stabilization by cobalamin 30 . In a more recent study, Gherasim et al showed that the MMACHC-R161Q mutant protein is associated with lower protein stability, decreased dealkylation, impaired glutathione binding and increased production of reactive oxygen species 28 . In this study, the authors compared two missense pathogenic variants involving this arginine residue; c.482G>A (p.Arg161Gln) and c.481C>G (p.Arg161Gly).…”

Section: Discussionmentioning

confidence: 99%

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Milder clinical and biochemical phenotypes associated with the c.482G > A (p.Arg161Gln) pathogenic variant in cobalamin C disease: Implications for management and screening

Almannai

Marom

Divin

et al. 2017

Molecular Genetics and Metabolism

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Introduction Cobalamin C disease is a multisystemic disease with variable manifestations and age of onset. Genotype-phenotype correlations are well-recognized in this disorder. Here, we present a large cohort of individuals with cobalamin C disease, several of whom are heterozygous for the c.482G>A pathogenic variant (p.Arg161Gln). We compared clinical characteristics of individuals with this pathogenic variant to those who do not have this variant. To our knowledge, this study represents the largest single cohort of individuals with the c.482G>A (p.Arg161Gln) pathogenic variant. Methods A retrospective chart review of 27 individuals from 21 families with cobalamin C disease who are followed at our facility was conducted. Results 13 individuals (48%) are compound heterozygous with the c.482G>A (p.Arg161Gln) on one allele and a second pathogenic variant on the other allele. Individuals with the c.482G>A (p.Arg161Gln) pathogenic variant had later onset of symptoms and easier metabolic control. Moreover, they had milder biochemical abnormalities at presentation which likely contributed to the observation that 4 individuals (31%) in this group were missed by newborn screening. Conclusion The c.482G>A (p.Arg161Gln) pathogenic variant is associated with milder disease. These individuals may not receive a timely diagnosis as they may not be identified on newborn screening or because of unrecognized, late onset symptoms. Despite the milder presentation, significant complications can occur, especially if treatment is delayed.

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Section: Discussionmentioning

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Milder clinical and biochemical phenotypes associated with the c.482G > A (p.Arg161Gln) pathogenic variant in cobalamin C disease: Implications for management and screening

Almannai

Marom

Divin

et al. 2017

Molecular Genetics and Metabolism

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“…The enzyme was first identified for its ability to reduce TNT (Bryant et al, 1991), but homologues have since been discovered with other activities including BluB and iodotyrosine deiodinase (Bang et al, 2012; Gherasim et al, 2015; Thomas et al, 2009; Yu et al, 2012). These raise the question of how the NR fold is able to support distinct activities in different exemplars, and of how NR is able to specialize in two-electron reduction yet apply this reactivity to diverse substrates.…”

Section: Introductionmentioning

confidence: 99%

Mechanism-Informed Refinement Reveals Altered Substrate-Binding Mode for Catalytically Competent Nitroreductase

et al. 2017

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Summary Nitroreductase from Enterobacter cloacae (NR) reduces diverse nitroaromatics including herbicides, explosives and prodrugs, and holds promise for bioremediation, prodrug activation and enzyme-assisted synthesis. We solved crystal structures of NR complexes with bound substrate or analog for each of its two half-reactions. We complemented these with kinetic isotope effect (KIE) measurements elucidating H-transfer steps essential to each half-reaction. KIEs indicate hydride transfer from NADH to the flavin consistent with our structure of NR with the NADH analog nicotinic acid adenine dinucleotide (NAAD). The KIE on reduction of p-nitrobenzoic acid (p-NBA) also indicates hydride transfer, and requires revision of prior computational mechanisms. Our mechanistic information provided a structural restraint for the orientation of bound substrate, placing the nitro group closer to the flavin N5 in the pocket that binds the amide of NADH. KIEs show that solvent provides a proton, enabling accommodation of different nitro group placements, consistent with NR’s broad repertoire.

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“…The thiol oxidase activity of hCblC is muted in contrast to that of ceCblC. Interestingly, two pathological mutants of hCblC, R161Q and R161G, exhibit elevated thiol oxidase activity, which, in turn, is correlated with increased oxidative stress that is associated with the CblC disorder (14).…”

mentioning

confidence: 99%

Coordination chemistry controls the thiol oxidase activity of the B12-trafficking protein CblC

Zhu

Shanmuganathan

Ruetz

et al. 2017

Journal of Biological Chemistry

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The cobalamin or B cofactor supports sulfur and one-carbon metabolism and the catabolism of odd-chain fatty acids, branched-chain amino acids, and cholesterol. CblC is a B-processing enzyme involved in an early cytoplasmic step in the cofactor-trafficking pathway. It catalyzes the glutathione (GSH)-dependent dealkylation of alkylcobalamins and the reductive decyanation of cyanocobalamin. CblC from (CblC) also exhibits a robust thiol oxidase activity, converting reduced GSH to oxidized GSSG with concomitant scrubbing of ambient dissolved O The mechanism of thiol oxidation catalyzed by CblC is not known. In this study, we demonstrate that novel coordination chemistry accessible toCblC-bound cobalamin supports its thiol oxidase activity via a glutathionyl-cobalamin intermediate. Deglutathionylation of glutathionyl-cobalamin by a second molecule of GSH yields GSSG. The crystal structure of CblC provides insights into how architectural differences at the α- and β-faces of cobalamin promote the thiol oxidase activity ofCblC but mute it in wild-type human CblC. The R161G and R161Q mutations in human CblC unmask its latent thiol oxidase activity and are correlated with increased cellular oxidative stress disease. In summary, we have uncovered key architectural features in the cobalamin-binding pocket that support unusual cob(II)alamin coordination chemistry and enable the thiol oxidase activity of CblC.

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Pathogenic Mutations Differentially Affect the Catalytic Activities of the Human B12-processing Chaperone CblC and Increase Futile Redox Cycling

Cited by 32 publications

References 51 publications

Milder clinical and biochemical phenotypes associated with the c.482G > A (p.Arg161Gln) pathogenic variant in cobalamin C disease: Implications for management and screening

Milder clinical and biochemical phenotypes associated with the c.482G > A (p.Arg161Gln) pathogenic variant in cobalamin C disease: Implications for management and screening

Mechanism-Informed Refinement Reveals Altered Substrate-Binding Mode for Catalytically Competent Nitroreductase

Coordination chemistry controls the thiol oxidase activity of the B12-trafficking protein CblC

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