Immunological homology among azoreductases from <i>Clostridium</i> and <i>Eubacterium</i> strains isolated from human intestinal microflora

Rafii, Fatemeh; Smith, David M.; Rw, Benson; Ce, Cerniglia

doi:10.1002/jobm.3620320204

Cited by 12 publications

(11 citation statements)

References 21 publications

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“…Intestinal microflora act in part as a barrier against bacterial pathogens and play an important role in metabolism of xenobiotics, drugs, and nutrients [2][3][4][5]. One example of the metabolism is the reduction of azo dyes to carcinogenic aromatic amines [6][7][8]. Azo dyes are compounds consisting of a diazotized amine coupled to an amine or a phenol, which contain one or more azo linkages.…”

Section: Introductionmentioning

confidence: 99%

The use of PCR to monitor the population abundance of six human intestinal bacterial species in an in vitro semicontinuous culture system

Wang

Cao

Campbell

et al. 1994

FEMS Microbiology Letters

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Six PCR primer sets complementary to the 16S rDNAs (rRNA genes) were developed and shown to be specific for the following anaerobic bacteria: Clostridium clostridiiforme, C. perfringens, C. leptum, Bacteroides vulgatus, B. distasonis, and B. thetaiotaomicron, respectively. These primers were used for PCR to detect and monitor the bacteria in a semicontinuous culture system designed to mimic intestinal microflora in the human gastrointestinal tract. Except for C. perfringens, the five species of Bacteroides and Clostridia present in the in vitro culture system were detected by the PCR, and the titers varied from 10 -2 to 10 6 dilutions. The role of azo dye reduction by these bacterial species in the system was examined and discussed.

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Section: Introductionmentioning

confidence: 99%

The use of PCR to monitor the population abundance of six human intestinal bacterial species in an in vitro semicontinuous culture system

Wang

Cao

Campbell

et al. 1994

FEMS Microbiology Letters

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“…The immunological cross-reactivity indicates that the azoreductase of C. perfringens shares structural similarities with the azoreductases from other species of anaerobic bacteria. These results suggest that the bacterial azoreductases tested may be considered a single related group of enzymes with regard to their function and antigenicity (11).…”

Section: Environmental Health Perspectivesmentioning

confidence: 87%

Reduction of Azo Dyes and Nitroaromatic Compounds by Bacterial Enzymes from the Human Intestinal Tract

Rafii¹,

Cerniglia²

1995

Environmental Health Perspectives

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Several anaerobic bacteria from the human intestinal tract are capable of reducing azo dyes and nitropolycyclic aromatic hydrocarbons to the corresponding aromatic amines with enzymes that have azoreductase and nitroreductase activities. The majority of bacteria with these activities belong to the genera Clostridium and Eubacterium. The azoreductases and nitroreductases from three Clostridium strains and one Eubacterium strain were studied. Both enzymes were produced constitutively in each of the bacteria; the enzymes from various bacteria had different electrophoretic mobilities. The azoreductases from all of the bacteria had immunological homology, as was evident from the cross-reactivity of an antibody raised against the azoreductase of C. perfringens with azoreductases from other bacteria. Comparison of azoreductases and nitroreductases showed that they both had identical electrophoretic mobilities on polyacrylamide gels and reacted with the antibody against the azoreductase from C. perfringens. Furthermore, the nitroaromatic compounds competitively inhibited the azoreductase activity. The data indicate that the reduction of both nitroaromatic compounds and azo dyes may be carried out by the same enzyme, which is possibly a flavin adenine dinucleotide dehydrogenase that is synthesized throughout the cell and not associated with any organized subcellular structure. -Environ Health Perspect 1 03(Suppl 5):1 7-19 (1995)

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“…After incubation for 3 hours at 42 °C, the phages were induced by overlaying the plates with nitrocellulose membranes saturated in 10 mM isopropyl b-D-thiogalactopyranoside (IPTG) and incubating at 37 °C for an additional 4 hours. The membranes then were washed and incubated at room temperature overnight with a rabbit anti-azoreductase antibody (RAFII et al 1992). Following three washes, the membranes were incubated with goat antirabbit IgG conjugated with peroxidase and then developed with a diaminobenzidine solution containing H 2 O 2 (RAFII et al 1993, HUYNH et al 1985.…”

Section: Dna Extractionmentioning

confidence: 99%

“…Several species of anaerobic bacteria that have azoreductase activity have previously been isolated from the human intestinal tract and shown to differ in the rates in which they reduce azo dyes (BRAGGER et al 1997, RAFII et al 1990. The azoreductases from these bacteria also vary in electrophoretic mobility but some of them have immunological homology with the C. perfringens azoreductase (RAFII et al 1992). Since no azoreductase genes from anaerobic bacteria have previously been cloned, the structural similarities of the azoreductase genes of different anaerobic bacteria are not known.…”

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confidence: 99%

Cloning and expression in Escherichia coli of an azoreductase gene from Clostridium perfringens and comparison with azoreductase genes from other bacteria

Rafii¹,

Coleman²

1999

J. Basic Microbiol.

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A genomic library of Clostridium perfringens ATCC 3626 was constructed in phage lambda gt11 and screened with an antibody against the C. perfringens azoreductase, which catalyzes the reduction of azo dyes to aromatic amines. A positive recombinant phage, containing a 3.8 kb DNA fragment insert was selected and purified. Lytic and lysogenic Escherichia coli cultures infected with the recombinant phage had higher azoreductase activity than cultures infected only with the vector lambda gt11. The 3.8 kb DNA fragment was amplified by PCR and found to hybridize with one band from C. perfringens DNA digested with EcoR1, indicating the presence of a single copy of the azoreductase gene. The fragment also hybridized with DNA from other azoreductase-producing Clostridium species, a Eubacterium sp., Enterobacter cloacae, Citrobacter amalonaticus and E. coli, but not with DNA from some other species of anaerobic bacteria capable of reducing azo dyes. The data indicate that the sequence of the azoreducatse gene of C. perfringens is conserved in some anaerobes and facultative anaerobes, but not in others, and that different types of azoreductase genes must be found in other anaerobic bacteria.

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Immunological homology among azoreductases from Clostridium and Eubacterium strains isolated from human intestinal microflora

Cited by 12 publications

References 21 publications

The use of PCR to monitor the population abundance of six human intestinal bacterial species in an in vitro semicontinuous culture system

The use of PCR to monitor the population abundance of six human intestinal bacterial species in an in vitro semicontinuous culture system

Reduction of Azo Dyes and Nitroaromatic Compounds by Bacterial Enzymes from the Human Intestinal Tract

Cloning and expression in Escherichia coli of an azoreductase gene from Clostridium perfringens and comparison with azoreductase genes from other bacteria

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