Acidic phospholipids inhibit the DNA-binding activity of DnaA protein, the initiator of chromosomal DNA replication in Escherichia coli

Masaki Makise, Shinji Mima, Takashi Katsu, Tomofusa Tsuchiya, Tohru Mizushima

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

In order to initiate chromosomal DNA replication in Escherichia coli, the DnaA protein must bind to both ATP and the origin of replication (oriC). Acidic phospholipids are known to inhibit DnaA binding to ATP, and here we examine the effects of various phospholipids on DnaA binding to oriC. Among the phospholipids in E. coli membrane, cardiolipin showed the strongest inhibition of DnaA binding to oriC. Synthetic phosphatidylglycerol containing unsaturated fatty acids inhibited binding more potently than did synthetic phosphatidylglycerol containing saturated fatty acids, suggesting that membrane fluidity is important. Thus, acidic phospholipids seem to inhibit DnaA binding to both oriC and adenine nucleotides in the same manner. Adenine nucleotides bound to DnaA did not affect the inhibitory effect of cardiolipin on DnaA binding to oriC. A mobility-shift assay revealed that acidic phospholipids inhibited formation of a DnaA-oriC complex containing several DnaA molecules. DNase I footprinting of DnaA binding to oriC showed that two DnaA binding sites (R2 and R3) were more sensitive to cardiolipin than other DnaA binding sites. Based on these in vitro data, the physiological relevance of this inhibitory effect of acidic phospholipids on DnaA binding to oriC is discussed.

Original languageEnglish
Pages (from-to)245-256
Number of pages12
JournalMolecular Microbiology
Volume46
Issue number1
DOIs
Publication statusPublished - 2002
Externally publishedYes

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Replication Origin
DNA Replication
Phospholipids
Escherichia coli
DNA
Cardiolipins
Proteins
Phosphatidylglycerols
Adenine Nucleotides
Adenosine Triphosphate
Binding Sites
Membrane Fluidity
Escherichia coli Proteins
Deoxyribonuclease I
Electrophoretic Mobility Shift Assay
Unsaturated Fatty Acids
Fatty Acids
Membranes

ASJC Scopus subject areas

  • Molecular Biology
  • Microbiology

Cite this

Acidic phospholipids inhibit the DNA-binding activity of DnaA protein, the initiator of chromosomal DNA replication in Escherichia coli. / Makise, Masaki; Mima, Shinji; Katsu, Takashi; Tsuchiya, Tomofusa; Mizushima, Tohru.

In: Molecular Microbiology, Vol. 46, No. 1, 2002, p. 245-256.

Research output: Contribution to journalArticle

Makise, Masaki ; Mima, Shinji ; Katsu, Takashi ; Tsuchiya, Tomofusa ; Mizushima, Tohru. / Acidic phospholipids inhibit the DNA-binding activity of DnaA protein, the initiator of chromosomal DNA replication in Escherichia coli. In: Molecular Microbiology. 2002 ; Vol. 46, No. 1. pp. 245-256.
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