A periplasmic, α-type carbonic anhydrase from Rhodopseudomonas palustris is essential for bicarbonate uptake

L. Puskás, M. Inui, K. Zahn, H. Yukawa

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

Intact cells of the purple non-sulfur bacterium Rhodopseudomonas palustris growing anaerobically, but not aerobically, contain carbonic anhydrase (CA) activity. The native enzyme was purified >2000-fold to apparent homogeneity and found to be a dimer with an estimated molecular mass of 54 kDa and a subunit molecular mass of 27 kDa. The CA gene (acaP) was cloned and its sequence revealed that it was homologous to α-type CAs. The upstream region of acaP was fused to the lacZ gene and β-galactosidase activity was measured under different growth conditions. Acetazolamide inhibited purified CA with an IC50 in the range of 10-8 M, and in the culture media concentrations as low as 30 μM inhibited phototrophic growth under anaerobic, light conditions when bicarbonate was used. An acaP::Kan(r) mutant strain was constructed by insertion of a kanamycin-resistance cassette and showed a growth pattern similar to wild-type cells grown in the presence of CA inhibitor. CO2 gas supplied as an inorganic carbon source reversed the effect of mutation or acetazolamide. CA activity measurements, fusion and Western blot experiments confirmed that CA is expressed under different anaerobic conditions independently of bicarbonate or CO2 and that there is no expression under aerobic conditions.

Original languageEnglish
Pages (from-to)2957-2966
Number of pages10
JournalMicrobiology
Volume146
Issue number11
Publication statusPublished - 2000

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Rhodopseudomonas
Carbonic Anhydrases
Bicarbonates
Acetazolamide
Phototrophic Processes
Chromatiaceae
Galactosidases
Kanamycin Resistance
Carbonic Anhydrase Inhibitors
Lac Operon
Growth
Inhibitory Concentration 50
Culture Media
Carbon
Gases
Western Blotting
Light
Mutation
Enzymes
Genes

Keywords

  • Carbonic anhydrase
  • Inorganic carbon uptake
  • Periplasmic enzyme
  • Rhodopseudomonas palustris

ASJC Scopus subject areas

  • Microbiology

Cite this

A periplasmic, α-type carbonic anhydrase from Rhodopseudomonas palustris is essential for bicarbonate uptake. / Puskás, L.; Inui, M.; Zahn, K.; Yukawa, H.

In: Microbiology, Vol. 146, No. 11, 2000, p. 2957-2966.

Research output: Contribution to journalArticle

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