Cloning and sequence analysis demonstrate the chromate reduction ability of a novel chromate reductase gene from Serratia sp.

Peng Deng,Qunhua Bai,Ying Wu,Xiaoqing Tan,Hong Xiao,Yan Jia

doi:10.3892/etm.2014.2148

Peng Deng, Qunhua Bai + Show 4 more

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https://doi.org/10.3892/etm.2014.2148

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Abstract

The ChrT gene encodes a chromate reductase enzyme which catalyzes the reduction of Cr(VI). The chromate reductase is also known as flavin mononucleotide (FMN) reductase (FMN_red). The aim of the present study was to clone the full-length ChrT DNA from Serratia sp. CQMUS2 and analyze the deduced amino acid sequence and three-dimensional structure. The putative ChrT gene fragment of Serratia sp. CQMUS2 was isolated by polymerase chain reaction (PCR), according to the known FMN_red gene sequence from Serratia sp. AS13. The flanking sequences of the ChrT gene were obtained by high efficiency TAIL-PCR, while the full-length gene of ChrT was cloned in Escherichia coli for subsequent sequencing. The nucleotide sequence of ChrT was submitted onto GenBank under the accession number, KF211434. Sequence analysis of the gene and amino acids was conducted using the Basic Local Alignment Search Tool, and open reading frame (ORF) analysis was performed using ORF Finder software. The ChrT gene was found to be an ORF of 567 bp that encodes a 188-amino acid enzyme with a calculated molecular weight of 20.4 kDa. In addition, the ChrT protein was hypothesized to be an NADPH-dependent FMN_red and a member of the flavodoxin-2 superfamily. The amino acid sequence of ChrT showed high sequence similarity to the FMN reductase genes of Klebsiella pneumonia and Raoultella ornithinolytica, which belong to the flavodoxin-2 superfamily. Furthermore, ChrT was shown to have a 85.6% similarity to the three-dimensional structure of Escherichia coli ChrR, sharing four common enzyme active sites for chromate reduction. Therefore, ChrT gene cloning and protein structure determination demonstrated the ability of the gene for chromate reduction. The results of the present study provide a basis for further studies on ChrT gene expression and protein function.

Highlights

Chromium is widely used as an important industrial material in leather tanning, dyeing, electroplating, pigment manufacturing and other industries
CQMUS2 cells was used as a template, an expected 305‐bp fragment of the ChrT gene was amplified using the Scfmn1‐F/R primers, which was subsequently sequenced (Fig. 1A)
The chromate reductase ChrT gene was cloned from Serratia sp

Summary

Introduction

Chromium is widely used as an important industrial material in leather tanning, dyeing, electroplating, pigment manufacturing and other industries. As a result of the secondary pollution and high cost, traditional, physical and chemical technologies cannot be extensively used to remedy the contaminated arable land and water. A number of microbial approaches to remedy chromium contamination have been investigated in attempts to overcome the problems [2,3,4]. Microbial bioremediation of chromium contamination is achieved mainly through two routes [5]. Previous studies have demonstrated that the ChrR enzyme from Pseudomonas putida [10], the YieF protein from Escherichia coli (E. coli) and the FMN_red from Pseudomonas aeruginosa (PAO1) have the ability to reduce Cr(VI) to Cr(III) [11,12], all of which are members of the FMN_red protein family

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