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. 1989 Sep;86(17):6612–6616. doi: 10.1073/pnas.86.17.6612

Genetically engineered mutant of the cyanobacterium Synechococcus PCC 7942 defective in nitrate transport

Tatsuo Omata *, Masayuki Ohmori †, Nobuyuki Arai *, Teruo Ogawa *
PMCID: PMC297894  PMID: 16594065

Abstract

Nitrate-grown cells of Synechococcus PCC 7942 (Anacystis nidulans R2) contain a 45-kDa protein as a major protein in the cytoplasmic membrane but ammonium-grown cells lack it. A mutant (M45) was constructed by inactivating the gene encoding the 45-kDa protein. M45 did not grow under low concentrations of nitrate but high concentrations of nitrate could support its growth, with the optimal concentration being 40-70 mM. The growth rate of M45 was as high as that of the wild-type cells when ammonium was the nitrogen source. The 45-kDa protein was absent in M45 irrespective of the growth conditions. The activities of nitrate and nitrite reductases were higher in M45 than in wild type. The rate of nitrate-dependent O2 evolution in wild type measured in the presence of L-methionine D,L-sulfoximine and D,L-glyceraldehyde showed saturation kinetics with respect to nitrate concentration in the external medium. The nitrate concentration required to produce half the maximal rate was 1 μM. In M45, the rate of nitrate-dependent O2 evolution was nearly zero at nitrate concentrations <1 mM and was linearly increased as the concentration increased. The presumed absence of nitrate transport in M45 demonstrated by these results suggested that the 45-kDa protein is a nitrate transporter.

Keywords: cytoplasmic membrane, 45-kDa protein, in vitro mutagenesis, nitrate reductase, ammonium-grown cell

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