Structural and functional study of ChuY from Escherichia coli strain CFT073

Kim, Hun; Chaurasia, Akhilesh Kumar; Kim, Truc; Choi, Joon‐Hwan; Ha, Soonhoi; Kim, Doyoun; Kim, Kyeong Kyu

doi:10.1016/j.bbrc.2016.12.008

Cited by 10 publications

(30 citation statements)

References 22 publications

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“…Finally, it is worth noting that when ChuY was deleted from pathogenic E. coli CFT073, a significant decrease in infection potential was observed. 23 Specifically, on the basis of cell counts, the ability of E. coli CFT073 to infect human embryonic kidney cells was significantly weakened in the absence of ChuY. A rational explanation for this observation is that the accumulation of heme or anaerobilin is toxic to the pathogen.…”

Section: Discussionmentioning

confidence: 99%

“…[26][27][28] Moreover, such kinetic behavior is inconsistent with ChuY functioning as a monomer. 23 In fact, equilibrium binding studies are the only way to provide evidence of site-to-site communication described by thermodynamic cooperativity. For example, the observation of a hyperbolic binding isotherm for binding of NADPH to ChuY would indicate a kinetic cooperative mechanism.…”

Section: Discussionmentioning

confidence: 99%

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Anaerobic Heme Degradation: ChuY Is an Anaerobilin Reductase That Exhibits Kinetic Cooperativity

LaMattina

Delrossi

et al. 2017

Biochemistry

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Heme catabolism is an important biochemical process that many bacterial pathogens utilize to acquire iron. However, tetrapyrrole catabolites can be reactive and often require further processing for transport out of the cell or conversion to another useful cofactor. In previous work, we presented in vitro evidence of an anaerobic heme degradation pathway in Escherichia coli O157:H7. Consistent with reactions that have been reported for other radical S-adenosyl-l-methionine methyltransferases, ChuW transfers a methyl group to heme by a radical-mediated mechanism and catalyzes the β-scission of the porphyrin macrocycle. This facilitates iron release and the production of a new linear tetrapyrrole termed "anaerobilin". In this work, we describe the structure and function of ChuY, an enzyme expressed downstream from chuW within the same heme utilization operon. ChuY is structurally similar to biliverdin reductase and forms a dimeric complex in solution that reduces anaerobilin to the product we have termed anaerorubin. Steady state analysis of ChuY exhibits kinetic cooperativity that is best explained by a random addition mechanism with a kinetically preferred path for initial reduced nicotinamide adenine dinucleotide phosphate binding.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Anaerobic Heme Degradation: ChuY Is an Anaerobilin Reductase That Exhibits Kinetic Cooperativity

LaMattina

Delrossi

et al. 2017

Biochemistry

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“…Preparation of polyclonal antibodies against rHutZ and immunoblot assays were performed as previously described [37]. Protease activity analysis of rHutZ was performed as reported by Kim et al [20]. Hemin-binding activity of rHutZ was evaluated as reported by Uchida et al [16].…”

Section: Methodsmentioning

confidence: 99%

“…hutWXZ and hugWXZ were considered necessary for obtaining iron from haem [17, 18]. In E. coli , a haem utilization gene cluster, chu , was identified that encodes a series of proteins, including ChuS, ChuA, ChuT, ChuW, ChuX, ChuY, and ChuU [19, 20]. ChuW and ChuX are homologous to HutW and HutX, which constitute the ChuW_HutW and ChuX_HutX superfamilies, respectively.…”

Section: Introductionmentioning

confidence: 99%

“…ChuY has high structural homology with human biliverdin and flavin reductase. It has been reported that ChuY has flavin mononucleotide (FMN) reductase activity, using NAD(P)H as a cofactor, and shows porphyrin ring binding affinity [19, 20]. Moreover, ChuY acts as a reductase in haem homeostasis to maintain the virulence potential of E. coli CFT073 [21].…”

Section: Introductionmentioning

confidence: 99%

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HutZ is required for biofilm formation and contributes to the pathogenicity of Edwardsiella piscicida

Shi

Fang

Huang

et al. 2019

Vet Res

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Edwardsiella piscicida is a severe fish pathogen. Haem utilization systems play an important role in bacterial adversity adaptation and pathogenicity. In this study, a speculative haem utilization protein, HutZ Ep , was characterized in E. piscicida. hutZ Ep is encoded with two other genes, hutW and hutX, in an operon that is similar to the haem utilization operon hutWXZ identified in V. cholerae. However, protein activity analysis showed that HutZ Ep is probably not related to hemin utilization. To explore the biological role of HutZ Ep , a markerless hutZ Ep in-frame mutant strain, TX01ΔhutZ, was constructed. Deletion of hutZ Ep did not significantly affect bacterial growth in normal medium, in iron-deficient conditions, or in the presence of haem but significantly retarded bacterial biofilm growth. The expression of known genes related to biofilm growth was not affected by hutZ Ep deletion, which indicated that HutZ Ep was probably a novel factor promoting biofilm formation in E. piscicida. Compared to the wild-type TX01, TX01ΔhutZ exhibited markedly compromised tolerance to acid stress and host serum stress. Pathogenicity analysis showed that inactivation of hutZ Ep significantly impaired the ability of E. piscicida to invade and reproduce in host cells and to infect host tissue. In contrast to TX01, TX01ΔhutZ was defective in blocking host macrophage activation. The expression of hutZ Ep was directly regulated by the ferric uptake regulator Fur. This study is the first functional characterization of HutZ in a fish pathogen, and these findings suggested that HutZ Ep is essential for E. piscicida biofilm formation and contributes to host infection.

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A new member of the flavodoxin superfamily from Fusobacterium nucleatum that functions in heme trafficking and reduction of anaerobilin

McGregor,

Chan,

Schroeder

et al. 2023

Journal of Biological Chemistry

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Structural and functional study of ChuY from Escherichia coli strain CFT073

Cited by 10 publications

References 22 publications

Anaerobic Heme Degradation: ChuY Is an Anaerobilin Reductase That Exhibits Kinetic Cooperativity

Anaerobic Heme Degradation: ChuY Is an Anaerobilin Reductase That Exhibits Kinetic Cooperativity

HutZ is required for biofilm formation and contributes to the pathogenicity of Edwardsiella piscicida

A new member of the flavodoxin superfamily from Fusobacterium nucleatum that functions in heme trafficking and reduction of anaerobilin

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