Amino-acid residues involved in the expression of the activity of Escherichia coli TolC

Hiroyasu Yamanaka, Naoyuki Morisada, Masaya Miyano, Hideaki Tsuge, Sumio Shinoda, Eizo Takahashi, Keinosuke Okamoto

Research output: Contribution to journalArticlepeer-review

11 Citations (Scopus)

Abstract

The Escherichia coli TolC, composed of 471 amino-acid residues, functions as a channel tunnel in the transport of various molecules across the outer membrane. We found previously that Leu-412, the 60th amino-acid residue from the carboxy terminal end, was crucial to the transport activity of TolC. Leu-412 is located in a domain which protrudes from the main body of TolC into the periplasm. Subsequent study indicated that the hydrophobicity generated by Leu-412 played an important role in the activity of TolC (H. Yamanaka, T. Nomura, N. Morisada, S. Shinoda, and K. Okamoto, Microb. Pathog. 33: 81-89, 2002). We predicted that other hydrophobic amino-acid residues around Leu-412 were also involved in the expression of the activity of TolC. To test this possibility, we substituted several hydrophobic residues around Leu-412, (Leu-3, Val-6, Leu-212, Leu-213, Leu-223, and Leu-224), with serine and examined the activity of these mutant TolCs. The result showed that Leu-3 is involved in the activity of TolC, but the other residues are not. The involvement of Leu-3 was confirmed by the residue deletion experiment. A subsequent point-mutational analysis of the residue showed that a hydrophobic side chain is required at position 3 for TolC to express its activity. As the distance between the α-carbons of Leu-3 and Leu-412 is just 7.45 Å, hydrophobic interaction between the two leucine residues might be involved in the activity of TolC.

Original languageEnglish
Pages (from-to)713-722
Number of pages10
JournalMICROBIOLOGY and IMMUNOLOGY
Volume48
Issue number10
DOIs
Publication statusPublished - 2004

Keywords

  • Escherichia coli
  • Heat-stable enterotoxin
  • Secretion
  • TolC

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Virology

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