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Biochemical Characterization of Escherichia coli FtsZ Protein Using Native 1 PAGE, Size-Exclusion Chromatography, Sedimentation Gradient, Chemical 2 Cross-Linking and Protein-Protein Interaction Prediction
Version 1
: Received: 23 August 2024 / Approved: 23 August 2024 / Online: 24 August 2024 (00:32:03 CEST)
How to cite:
Araujo, N.; Veloso, M.; Pouchucq, L. Biochemical Characterization of Escherichia coli FtsZ Protein Using Native 1 PAGE, Size-Exclusion Chromatography, Sedimentation Gradient, Chemical 2 Cross-Linking and Protein-Protein Interaction Prediction. Preprints2024, 2024081748. https://doi.org/10.20944/preprints202408.1748.v1
Araujo, N.; Veloso, M.; Pouchucq, L. Biochemical Characterization of Escherichia coli FtsZ Protein Using Native 1 PAGE, Size-Exclusion Chromatography, Sedimentation Gradient, Chemical 2 Cross-Linking and Protein-Protein Interaction Prediction. Preprints 2024, 2024081748. https://doi.org/10.20944/preprints202408.1748.v1
Araujo, N.; Veloso, M.; Pouchucq, L. Biochemical Characterization of Escherichia coli FtsZ Protein Using Native 1 PAGE, Size-Exclusion Chromatography, Sedimentation Gradient, Chemical 2 Cross-Linking and Protein-Protein Interaction Prediction. Preprints2024, 2024081748. https://doi.org/10.20944/preprints202408.1748.v1
APA Style
Araujo, N., Veloso, M., & Pouchucq, L. (2024). Biochemical Characterization of Escherichia coli FtsZ Protein Using Native 1 PAGE, Size-Exclusion Chromatography, Sedimentation Gradient, Chemical 2 Cross-Linking and Protein-Protein Interaction Prediction. Preprints. https://doi.org/10.20944/preprints202408.1748.v1
Chicago/Turabian Style
Araujo, N., Marcelo Veloso and Luis Pouchucq. 2024 "Biochemical Characterization of Escherichia coli FtsZ Protein Using Native 1 PAGE, Size-Exclusion Chromatography, Sedimentation Gradient, Chemical 2 Cross-Linking and Protein-Protein Interaction Prediction" Preprints. https://doi.org/10.20944/preprints202408.1748.v1
Abstract
FtsZ is a bacterial protein that plays a crucial role in cytokinesis by forming the Z-ring. This ring acts as a scaffold to recruit other division proteins and guide the synthesis of septal peptidoglycan, which leads to cell constriction. In its native state, the FtsZ protein from Escherichia coli (EcFtsZ) exists as a multi-oligomer comprising dimers, trimers, tetramers, and hexamers in dynamic equilibrium of self-association and dissociation is dependent on protein concentration. This study employed classical biochemical techniques, including native PAGE, size-exclusion chromatography, sucrose gradient, and chemical cross-linking with formaldehyde, to characterize the trimeric state of EcFtsZ. The results of size-exclusion chromatography demonstrate that the trimer of EcFtsZ has a mass of 131 kDa and an experimental friction quotient (Rs/Rmin) of 1.9, which is very similar to the theoretical friction quotient (fn/f1) of 1.8 for a linear trimer. Cross-linking EcFtsZ with formaldehyde resulted in the occurrence of a polypeptide band of 128 kDa, which was recognized by anti-FtsZ antibodies. The result of the sedimentation of EcFtsZ cross-linking with formaldehyde returned a value (Smax/S20,w) equal to 1.9. This experimental value was found to be equal to the theoretical calculation of the sedimentation quotient (Sn/S1) for a lateral trimer. These findings indicate that at micromolar concentrations, the EcFtsZ trimer may assume a linear or lateral structure in the protein. Finally, the protein-protein interaction prediction program (PEPPI) indicated the potential for EcFtsZ to form two protein-protein contacts in its C-terminal domain, which may be involved in trimer formation.
Biology and Life Sciences, Biochemistry and Molecular Biology
Copyright:
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
