Partial Purification and Characterization of Chromate Reductase of a Novel<i>Ochrobactrum</i>sp. Strain Cr-B4

Hora, Anuradha; Shetty, Vidya K.

doi:10.1080/10826068.2014.952385

Cited by 22 publications

(12 citation statements)

References 36 publications

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“…Although Hora described supplementation of Fe (III) substantially enhanced the activity of chromate reductase purified from Ochrobactrum sp . Cr-B4 35 , Fe (III) has no significant effect on NfoR activity (Fig. 4 ).…”

Section: Resultsmentioning

confidence: 87%

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Copper (II) binding of NAD(P)H- flavin oxidoreductase (NfoR) enhances its Cr (VI)-reducing ability

Han

Ling

Zhou

et al. 2017

Sci Rep

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Microbes can reduce hexavalent chromium Cr (VI) to the less toxic and soluble trivalent Cr (III). Copper stimulates microbial reduction of Cr (VI) by the Bacillus, Ochrobactrum, and Gluconobacter species; however, the mechanism remains unclear. In our study, the rate of Cr (VI) reduction by Staphylococcus aureus LZ-01 was increased by 210 % when supplemented with 60 μM Cu (II). A putative NAD(P)H-flavin oxidoreductase gene (nfoR) was upregulated under Cr (VI) stress. NfoR-knockout mutant displayed impaired reduction of Cr (VI) and Cu (II)-enhanced Cr (VI) reduction by nfoR isogenic mutant was attenuated in the presence of Cu (II). In vitro tests showed an increased V max value of 25.22 μM min−1 mg−1 NfoR in the presence of Cu (II). Together, these results indicate that NfoR is responsible for Cu (II) enhancement. Isothermal titration calorimetry (ITC) assays confirmed the interaction of NfoR with Cu (II) at the dissociation constant of 85.5 μM. Site-directed mutagenesis indicates that His100, His128, and Met165 residues may be important for Cu (II) binding, while Cys163 is necessary for the FMN binding of NfoR. These findings show that Cu (II)-enhanced NfoR belongs to a new branch of Cr (VI) reductases and profoundly influences Cr (VI) reduction.

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Section: Resultsmentioning

confidence: 87%

“…Cu (II) and Fe (III) substantially enhance the activity of chromate reductase purified from Ochrobactrum sp . Cr-B4 35 . Similar phenomena are observed in a number of strains within the Bacillus genus 36 .…”

Section: Introductionmentioning

confidence: 99%

Copper (II) binding of NAD(P)H- flavin oxidoreductase (NfoR) enhances its Cr (VI)-reducing ability

Han

Ling

Zhou

et al. 2017

Sci Rep

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“…Nevertheless, typical for soil bacteria, some genes were identified on the POA1180 genome, which could be beneficial for its host. Resistance against chromate has been found in several bacteria including Ochrobactrum (Branco and Morais, 2013 ; Hora and Shetty, 2015 ), but to the best of our knowledge, the respective genes have yet not been identified on the genome of a temperate phage. The same holds true for the sulfate permease gene which may improve the supply of the host with sulfate (Pilsyk and Paszewski, 2009 ).…”

Section: Discussionmentioning

confidence: 97%

Prevalence, Host Range, and Comparative Genomic Analysis of Temperate Ochrobactrum Phages

et al. 2017

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Ochrobactrum and Brucella are closely related bacteria that populate different habitats and differ in their pathogenic properties. Only little is known about mobile genetic elements in these genera which might be important for survival and virulence. Previous studies on Brucella lysogeny indicated that active phages are rare in this genus. To gain insight into the presence and nature of prophages in Ochrobactrum, temperate phages were isolated from various species and characterized in detail. In silico analyses disclosed numerous prophages in published Ochrobactrum genomes. Induction experiments showed that Ochrobactrum prophages can be induced by various stress factors and that some strains released phage particles even under non-induced conditions. Sixty percent of lysates prepared from 125 strains revealed lytic activity. The host range and DNA similarities of 19 phages belonging to the families Myoviridae, Siphoviridae, or Podoviridae were determined suggesting that they are highly diverse. Some phages showed relationship to the temperate Brucella inopinata phage BiPB01. The genomic sequences of the myovirus POA1180 (41,655 bp) and podovirus POI1126 (60,065 bp) were analyzed. Phage POA1180 is very similar to a prophage recently identified in a Brucella strain isolated from an exotic frog. The POA1180 genome contains genes which may confer resistance to chromate and the ability to take up sulfate. Phage POI1126 is related to podoviruses of Sinorhizobium meliloti (PCB5), Erwinia pyrifoliae (Pep14), and Burkholderia cenocepacia (BcepIL02) and almost identical to an unnamed plasmid of the Ochrobactrum intermedium strain LMG 3301. Further experiments revealed that the POI1126 prophage indeed replicates as an extrachromosomal element. The data demonstrate for the first time that active prophages are common in Ochrobactrum and suggest that atypical brucellae also may be a reservoir for temperate phages.

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“…Although Cu(II) stimulates the activity of chromate reductase from Ochrobactrum sp . Cr‐B4 (Hora and Shetty, 2015), the detailed information on Cu(II)‐stimulated chromate reductase activity is unavailable. Thus, catalytic mechanisms are fundamental for understanding structure–function relationships.…”

Section: Introductionmentioning

confidence: 99%

Cu(II) nonspecifically binding chromate reductase NfoR promotes Cr(VI) reduction

Han

Zheng

Zhou

et al. 2020

Environmental Microbiology

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Summary Cu(II)‐enhanced microbial Cr(VI) reduction is common in the environment, yet its mechanism is unknown. The specific activity of chromate reductase, NfoR, from Staphylococcus aureus sp. LZ‐01 was augmented 1.5‐fold by Cu(II). Isothermal titration calorimetry and spectral data show that Cu(II) binds to NfoR nonspecifically. Further, Cu(II) stimulates the nitrobenzene reduction of NfoR, indicating that Cu(II) promotes electron transfer. The crystal structure of NfoR in complex with CuSO4 (1.46 Å) was determined. The overall structure of NfoR‐Cu(II) complex is a dimer that covalently binds with FMN and Cu(II)‐binding pocket is located at the interface of the NfoR dimer. Structural superposition revealed that NfoR resembles the structure of class II chromate reductase. Site‐directed mutagenesis revealed that Leu46 and Phe123 were involved in NADH binding, whereas Trp70 and Ser45 were the key residues for nitrobenzene binding. Furthermore, His100 and Asp171 were preferential affinity sites for Cu(II) and that Cys163 is an active site for FMN binding. Attenuation reductase activity in C163S can be partially restored to 54% wild type by increasing Cu(II) concentration. Partial restoration indicates dual‐channel electron transfer of NfoR via Cu(II) and FMN. We propose a catalytic mechanism for Cu(II)‐enhanced NfoR activity in which Cu(I) is formed transiently. Together, the current results provide an insight on Cu (II)‐induced enhancement and benefit of Cr(VI) bioremediation.

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Partial Purification and Characterization of Chromate Reductase of a NovelOchrobactrumsp. Strain Cr-B4

Cited by 22 publications

References 36 publications

Copper (II) binding of NAD(P)H- flavin oxidoreductase (NfoR) enhances its Cr (VI)-reducing ability

Copper (II) binding of NAD(P)H- flavin oxidoreductase (NfoR) enhances its Cr (VI)-reducing ability

Prevalence, Host Range, and Comparative Genomic Analysis of Temperate Ochrobactrum Phages

Cu(II) nonspecifically binding chromate reductase NfoR promotes Cr(VI) reduction

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