Aerobic-Type Ribonucleotide Reductase in the Anaerobe Bacteroides fragilis

ECU Author/Contributor (non-ECU co-authors, if there are any, appear on document)

Edson R. Rocha (Creator)

Darren Smalley (Creator)

C. Jeffrey Smith (Creator)

Institution

East Carolina University (ECU )

Web Site: http://www.ecu.edu/lib/

Abstract: Bacteroides fragilis a component of the normal intestinal flora is an obligate anaerobe capable of long-term survival in the presence of air. Survival is attributed to an elaborate oxidative stress response that controls the induction of more than 28 peptides but there is limited knowledge concerning the identities of these peptides. In this report RNA fingerprinting by arbitrarily primed PCR identified five new genes whose expression increased following exposure to O2. Nucleotide sequence analysis of the cloned genes indicated that they encoded an outer membrane protein an aspartate decarboxylase an efflux pump heat shock protein HtpG and an NrdA ortholog constituting the large subunit of a class Ia ribonucleotide reductase (RRase). Attention was focused on the nrdA gene since class I RRases are obligate aerobic enzymes catalyzing the reduction of ribonucleoside 5'-diphosphates by a mechanism that requires molecular oxygen for activity. Sequence analysis of the nrd locus showed that two genes nrdA and nrdB are located in the same orientation in a 4.5-kb region. Northern hybridization and primer extension experiments confirmed induction of the genes by O2 and suggested they are an operon. The B. fragilis nrdA and nrdB genes were overexpressed in Escherichia coli and CDP reductase assays confirmed that they encoded an active enzyme. The enzyme activity was inhibited by hydroxyurea and ATP was shown to be a positive effector of CDP reductase activity while dATP was an inhibitor indicating that the enzyme was a class Ia RRase. A nrdA mutant was viable under anaerobic conditions but had decreased survival following exposure to O2 and it could not rapidly resume growth after O2 treatment. The results presented indicate that during aerobic conditions B. fragilis NrdAB may have a role in maintaining deoxyribonucleotide pools for DNA repair and growth recovery. Originally published Journal of Bacteriology Vol. 184 No. 4 Feb 2002

Additional Information

Publication

Other

Journal of Bacteriology. 184:4(February 2002) p. 895-903.

Language: English

Date: 2011

Keywords

Bacteroides fragilis, Ribonucleotide Reductases, aerobic environment

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