Analysis of genes involved in nitrate reduction in Clostridium perfringens

Katsuyo Fujinaga, Yuki Taniguchi, Yezhou Sun, Seiichi Katayama, Junzaburo Minami, Osamu Matsushita, Akinobu Okabe

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

We have conducted the genetic analysis of fermentative nitrate reduction in Clostridium perfringens, a strict anaerobic bacterium. Nitrate reductase (NarA) was purified from the cytoplasmic fraction of the organism. Using a degenerate primer designed from its N-terminal amino acid sequence, a 9.5 kb fragment containing seven ORFs was cloned. The molecular mass and N-terminal amino acid sequence predicted from the nucleotide sequence of ORF 4 coincided with those determined for the purified NarA, indicating that ORF 4 corresponds to a narA gene. ORFs 5 and 6 encode a 15.4 kDa ferredoxin-like protein containing four iron-sulfur clusters and a 45 kDa protein homologous to NADH oxidase, respectively. Analyses involving primer extension and Northern blotting revealed that these three ORFs are transcribed as a polycistronic message. The ORF 5- and ORF 6-encoded proteins were shown by immunablotting to be synthesized by cells grown in the presence of nitrate. Thus, these two proteins are likely to function as electron-transfer components in nitrate reduction in C. perfringens. The 9.5 kb fragment and a downstream region of 6.1 kb do not contain any genes involved in nitrate uptake or nitrite reduction. Instead, all 5 ORFs downstream of ORF 6 are homologous to genes reported for molybdopterin biosynthesis, unlike the genomic organization already determined for the respiratory and assimilatory nitrate-reduction systems. The evolutionary relationships between these two nitrate-reduction systems and the fermentative one based on the results of comparative genetic analysis are discussed.

Original languageEnglish
Pages (from-to)3377-3387
Number of pages11
JournalMicrobiology
Volume145
Issue number12
DOIs
Publication statusPublished - Dec 1999
Externally publishedYes

Keywords

  • Clostridium perfringens
  • Ferredoxin-like protein
  • Molybdopterin guanine dinucleotide
  • NADH oxidase
  • Nitrate reductase

ASJC Scopus subject areas

  • Microbiology

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