Interaction of the transmembrane domain of lysis protein E from bacteriophage phiX174 with bacterial translocase MraY and peptidyl-prolyl isomerase SlyD

Sharon Mendel, Joanne M Holbourn, James A Schouten, Timothy D H Bugg

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

The molecular target for the bacteriolytic E protein from bacteriophage X174, responsible for host cell lysis, is known to be the enzyme phospho-MurNAc-pentapeptide translocase (MraY), an integral membrane protein involved in bacterial cell wall peptidoglycan biosynthesis, with an essential role being played by peptidyl-prolyl isomerase SlyD. A synthetic 37 aa peptide E(pep), containing the N-terminal transmembrane alpha-helix of E, was found to be bacteriolytic against Bacillus licheniformis, and inhibited membrane-bound MraY. The solution conformation of E(pep) was found by circular dichroism (CD) spectroscopy to be 100 % alpha-helical. No change in the CD spectrum was observed upon addition of purified Escherichia coli SlyD, implying that SlyD does not catalyse prolyl isomerization upon E. However, E(pep) was found to be a potent inhibitor of SlyD-catalysed peptidyl-prolyl isomerization (IC(50) 0.15 microM), implying a strong interaction between E and SlyD. E(pep) was found to inhibit E. coli MraY activity when assayed in membranes (IC(50) 0.8 microM); however, no inhibition of solubilized MraY was observed, unlike nucleoside natural product inhibitor tunicamycin. These results imply that the interaction of E with MraY is not at the MraY active site, and suggest that a protein-protein interaction is formed between E and MraY at a site within the transmembrane region.

Original languageEnglish
Pages (from-to)2959-2967
Number of pages9
Journal BMC Microbiology
Volume152
Issue numberPt 10
Early online date1 Oct 2006
DOIs
Publication statusPublished - Oct 2006

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Peptidyl Transferases
Peptidylprolyl Isomerase
Circular Dichroism
Bacteriophages
Escherichia coli
Tunicamycin
Membranes
Peptidoglycan
Biological Products
Nucleosides
Cell Wall
Catalytic Domain
Spectrum Analysis
Membrane Proteins
Proteins
Enzymes
Protein Domains
alpha-Helical Protein Conformation
peptide E (adrenal medulla)
bacteriophage X174 E protein

Keywords

  • Anti-Bacterial Agents
  • Bacillus
  • Bacterial Proteins
  • Bacteriolysis
  • Circular Dichroism
  • Escherichia coli Proteins
  • Models, Biological
  • Peptides
  • Peptidylprolyl Isomerase
  • Protein Binding
  • Protein Interaction Mapping
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Transferases
  • Viral Proteins
  • Journal Article
  • Research Support, Non-U.S. Gov't

Cite this

Interaction of the transmembrane domain of lysis protein E from bacteriophage phiX174 with bacterial translocase MraY and peptidyl-prolyl isomerase SlyD. / Mendel, Sharon; Holbourn, Joanne M; Schouten, James A; Bugg, Timothy D H.

In: BMC Microbiology , Vol. 152, No. Pt 10, 10.2006, p. 2959-2967.

Research output: Contribution to journalArticle

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abstract = "The molecular target for the bacteriolytic E protein from bacteriophage X174, responsible for host cell lysis, is known to be the enzyme phospho-MurNAc-pentapeptide translocase (MraY), an integral membrane protein involved in bacterial cell wall peptidoglycan biosynthesis, with an essential role being played by peptidyl-prolyl isomerase SlyD. A synthetic 37 aa peptide E(pep), containing the N-terminal transmembrane alpha-helix of E, was found to be bacteriolytic against Bacillus licheniformis, and inhibited membrane-bound MraY. The solution conformation of E(pep) was found by circular dichroism (CD) spectroscopy to be 100 {\%} alpha-helical. No change in the CD spectrum was observed upon addition of purified Escherichia coli SlyD, implying that SlyD does not catalyse prolyl isomerization upon E. However, E(pep) was found to be a potent inhibitor of SlyD-catalysed peptidyl-prolyl isomerization (IC(50) 0.15 microM), implying a strong interaction between E and SlyD. E(pep) was found to inhibit E. coli MraY activity when assayed in membranes (IC(50) 0.8 microM); however, no inhibition of solubilized MraY was observed, unlike nucleoside natural product inhibitor tunicamycin. These results imply that the interaction of E with MraY is not at the MraY active site, and suggest that a protein-protein interaction is formed between E and MraY at a site within the transmembrane region.",
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T1 - Interaction of the transmembrane domain of lysis protein E from bacteriophage phiX174 with bacterial translocase MraY and peptidyl-prolyl isomerase SlyD

AU - Mendel, Sharon

AU - Holbourn, Joanne M

AU - Schouten, James A

AU - Bugg, Timothy D H

PY - 2006/10

Y1 - 2006/10

N2 - The molecular target for the bacteriolytic E protein from bacteriophage X174, responsible for host cell lysis, is known to be the enzyme phospho-MurNAc-pentapeptide translocase (MraY), an integral membrane protein involved in bacterial cell wall peptidoglycan biosynthesis, with an essential role being played by peptidyl-prolyl isomerase SlyD. A synthetic 37 aa peptide E(pep), containing the N-terminal transmembrane alpha-helix of E, was found to be bacteriolytic against Bacillus licheniformis, and inhibited membrane-bound MraY. The solution conformation of E(pep) was found by circular dichroism (CD) spectroscopy to be 100 % alpha-helical. No change in the CD spectrum was observed upon addition of purified Escherichia coli SlyD, implying that SlyD does not catalyse prolyl isomerization upon E. However, E(pep) was found to be a potent inhibitor of SlyD-catalysed peptidyl-prolyl isomerization (IC(50) 0.15 microM), implying a strong interaction between E and SlyD. E(pep) was found to inhibit E. coli MraY activity when assayed in membranes (IC(50) 0.8 microM); however, no inhibition of solubilized MraY was observed, unlike nucleoside natural product inhibitor tunicamycin. These results imply that the interaction of E with MraY is not at the MraY active site, and suggest that a protein-protein interaction is formed between E and MraY at a site within the transmembrane region.

AB - The molecular target for the bacteriolytic E protein from bacteriophage X174, responsible for host cell lysis, is known to be the enzyme phospho-MurNAc-pentapeptide translocase (MraY), an integral membrane protein involved in bacterial cell wall peptidoglycan biosynthesis, with an essential role being played by peptidyl-prolyl isomerase SlyD. A synthetic 37 aa peptide E(pep), containing the N-terminal transmembrane alpha-helix of E, was found to be bacteriolytic against Bacillus licheniformis, and inhibited membrane-bound MraY. The solution conformation of E(pep) was found by circular dichroism (CD) spectroscopy to be 100 % alpha-helical. No change in the CD spectrum was observed upon addition of purified Escherichia coli SlyD, implying that SlyD does not catalyse prolyl isomerization upon E. However, E(pep) was found to be a potent inhibitor of SlyD-catalysed peptidyl-prolyl isomerization (IC(50) 0.15 microM), implying a strong interaction between E and SlyD. E(pep) was found to inhibit E. coli MraY activity when assayed in membranes (IC(50) 0.8 microM); however, no inhibition of solubilized MraY was observed, unlike nucleoside natural product inhibitor tunicamycin. These results imply that the interaction of E with MraY is not at the MraY active site, and suggest that a protein-protein interaction is formed between E and MraY at a site within the transmembrane region.

KW - Anti-Bacterial Agents

KW - Bacillus

KW - Bacterial Proteins

KW - Bacteriolysis

KW - Circular Dichroism

KW - Escherichia coli Proteins

KW - Models, Biological

KW - Peptides

KW - Peptidylprolyl Isomerase

KW - Protein Binding

KW - Protein Interaction Mapping

KW - Protein Structure, Secondary

KW - Protein Structure, Tertiary

KW - Transferases

KW - Viral Proteins

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

U2 - 10.1099/mic.0.28776-0

DO - 10.1099/mic.0.28776-0

M3 - Article

VL - 152

SP - 2959

EP - 2967

JO - BMC Microbiology

JF - BMC Microbiology

SN - 1471-2180

IS - Pt 10

ER -