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Acidic phospholipids inhibit the DNA‐binding activity of DnaA protein, the initiator of chromosomal DNA replication in Escherichia coli
Author(s) -
Makise Masaki,
Mima Shinji,
Katsu Takashi,
Tsuchiya Tomofusa,
Mizushima Tohru
Publication year - 2002
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.2002.03161.x
Subject(s) - dnaa , cardiolipin , phosphatidylglycerol , biology , dnab helicase , biochemistry , dna , binding site , dna replication , dna binding protein , prokaryotic dna replication , origin of replication , phospholipid , helicase , rna , gene , membrane , transcription factor , phosphatidylcholine
Summary 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 re‐vealed 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.