Dimerization of signalling modules of the EvgAS and BvgAS phosphorelay systems

Biophysical and biochemical properties of signalling proteins or domains derived from the unorthodox EvgAS and BvgAS two-component phosphorelay systems of Escherichia coli and Bordetella pertussis were investigated. Oligomerization of the effector proteins EvgA and BvgA and of truncated EvgS and Bvg...

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Bibliographic Details
Published in:Biochimica et biophysica acta 2000-05, Vol.1478 (2), p.341-354
Main Authors: Perraud, Anne-Laure, Rippe, Karsten, Bantscheff, Marcus, Glocker, Michael, Lucassen, Magnus, Jung, Kirsten, Sebald, Walter, Weiss, Verena, Gross, Roy
Format: Article
Language:English
Subjects:
Analytical ultracentrifugation
Bordetella pertussis - chemistry
BvgAS
Chromatography, Gel
Dimerization
Escherichia coli - chemistry
EvgAS
Histidine Kinase
Mass spectrometry
Molecular Weight
Phosphorelay
Phosphorylation
Protein Kinases - chemistry
Protein Sorting Signals - chemistry
Signal Transduction
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Surface Plasmon Resonance
Ultracentrifugation
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Summary:Biophysical and biochemical properties of signalling proteins or domains derived from the unorthodox EvgAS and BvgAS two-component phosphorelay systems of Escherichia coli and Bordetella pertussis were investigated. Oligomerization of the effector proteins EvgA and BvgA and of truncated EvgS and BvgS derived signalling proteins containing the receiver and histidine containing phosphotransfer (HPt) domains or comprising only the HPt domains were characterized by native gel electrophoresis, gel permeation experiments and analytical ultracentrifugation. The results obtained by the different methods are consistent with non-phosphorylated EvgA and BvgA proteins being dimers in solution with a dissociation constant significantly below 1 μM. In contrast, all sensor derived domains of EvgS and BvgS were observed to be monomers in vitro. No indications for a phosphorylation induced stimulation of oligomerization of the C-terminal histidine kinase domains could be detected. In agreement with these data, surface plasmon resonance studies revealed a 2:1 stoichiometry in the interaction of EvgA with the immobilized EvgS HPt domain and an affinity constant of 1.24×10 6 M −1.
ISSN:0167-4838
0006-3002
1879-2588
DOI:10.1016/S0167-4838(00)00052-2