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Mapping the Contact Sites of the Escherichia coli Division-Initiating Proteins FtsZ and ZapA by BAMG Cross-Linking and Site-Directed Mutagenesis

1
Mass Spectrometry of Biomacromolecules, Swammerdam Institute for Life Sciences, Faculty of Science, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands
2
Bacterial Cell Biology and Physiology, Swammerdam Institute for Life Sciences, Faculty of Science, University of Amsterdam, Science Park 904, 1098 XH Amsterdam, The Netherlands
3
Computational Structural Biology, Faculty of Science-Chemistry, University of Utrecht, Padualaan 83584CH Utrecht, The Netherlands
4
Faculty of Medical Technology, Prince of Songkla University, Songkhla 90110, Thailand
*
Authors to whom correspondence should be addressed.
Int. J. Mol. Sci. 2018, 19(10), 2928; https://doi.org/10.3390/ijms19102928
Received: 23 August 2018 / Revised: 13 September 2018 / Accepted: 19 September 2018 / Published: 26 September 2018
(This article belongs to the Special Issue Regulatory Mechanisms of Tubulin-Like Proteins)
Cell division in bacteria is initiated by the polymerization of FtsZ at midcell in a ring-like structure called the Z-ring. ZapA and other proteins assist Z-ring formation and ZapA binds ZapB, which senses the presence of the nucleoids. The FtsZ–ZapA binding interface was analyzed by chemical cross-linking mass spectrometry (CXMS) under in vitro FtsZ-polymerizing conditions in the presence of GTP. Amino acids residue K42 from ZapA was cross-linked to amino acid residues K51 and K66 from FtsZ, close to the interphase between FtsZ molecules in protofilaments. Five different cross-links confirmed the tetrameric structure of ZapA. A number of FtsZ cross-links suggests that its C-terminal domain of 55 residues, thought to be largely disordered, has a limited freedom to move in space. Site-directed mutagenesis of ZapA reveals an interaction site in the globular head of the protein close to K42. Using the information on the cross-links and the mutants that lost the ability to interact with FtsZ, a model of the FtsZ protofilament–ZapA tetramer complex was obtained by information-driven docking with the HADDOCK2.2 webserver. View Full-Text
Keywords: cell division; Z associated protein A (ZapA); Filamenting temperature sensitive Z (FtsZ); quadrupole time of flight mass spectrometer (QTOF); Fourier-Transform Ion Cyclotron Resonance mass spectrometry(FTICR); 1,4-bis(succimidyl)-3-azidomethylglutarate (BAMG) cell division; Z associated protein A (ZapA); Filamenting temperature sensitive Z (FtsZ); quadrupole time of flight mass spectrometer (QTOF); Fourier-Transform Ion Cyclotron Resonance mass spectrometry(FTICR); 1,4-bis(succimidyl)-3-azidomethylglutarate (BAMG)
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Roseboom, W.; Nazir, M.G.; Meiresonne, N.Y.; Mohammadi, T.; Verheul, J.; Buncherd, H.; Bonvin, A.M.J.J.; De Koning, L.J.; De Koster, C.G.; De Jong, L.; Den Blaauwen, T. Mapping the Contact Sites of the Escherichia coli Division-Initiating Proteins FtsZ and ZapA by BAMG Cross-Linking and Site-Directed Mutagenesis. Int. J. Mol. Sci. 2018, 19, 2928.

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