Regulation of signaling directionality revealed by 3D snapshots of a kinase:regulator complex in action

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Abstract

Two-component systems (TCS) are protein machineries that enable cells to respond to input signals. Histidine kinases (HK) are the sensory component, transferring information toward downstream response regulators (RR). HKs transfer phosphoryl groups to their specific RRs, but also dephosphorylate them, overall ensuring proper signaling. The mechanisms by which HKs discriminate between such disparate directions, are yet unknown. We now disclose crystal structures of the HK:RR complex DesK:DesR from Bacillus subtilis, comprising snapshots of the phosphotransfer and the dephosphorylation reactions. The HK dictates the reactional outcome through conformational rearrangements that include the reactive histidine. The phosphotransfer center is asymmetric, poised for dissociative nucleophilic substitution. The structural bases of HK phosphatase/phosphotransferase control are uncovered, and the unexpected discovery of a dissociative reactional center, sheds light on the evolution of TCS phosphotransfer reversibility. Our findings should be applicable to a broad range of signaling systems and instrumental in synthetic TCS rewiring.

Data availability

The following data sets were generated

1. Trajtenberg F
2. Buschiazzo A
(2016) Crystal structure of phosphorylated DesKC
Publicly available at the RCSB Protein Data Bank (accession no: 5IUM).

http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5IUM
(2016) Crystal structure of the DesK-DesR complex in the phosphatase state
Publicly available at the RCSB Protein Data Bank (accession no: 5IUN).

http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5IUN
(2016) Crystal structure of the DesK-DesR complex in the phosphotransfer state with low Mg2+ (20 mM)
Publicly available at the RCSB Protein Data Bank (accession no: 5IUJ).

http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5IUJ
(2016) Crystal structure of the DesK-DesR complex in the phosphotransfer state with high Mg2+ (150 mM)
Publicly available at the RCSB Protein Data Bank (accession no: 5IUK).

http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5IUK
(2016) Crystal structure of the DesK-DesR complex in the phosphotransfer state with high Mg2+ (150 mM) and BeF3
Publicly available at the RCSB Protein Data Bank (accession no: 5IUL).

http://www.rcsb.org/pdb/search/structidSearch.do?structureId=5IUL

Article and author information

Author details

Felipe Trajtenberg

Laboratory of Molecular and Structural Microbiology, Institut Pasteur de Montevideo, Montevideo, Uruguay

Competing interests
The authors declare that no competing interests exist.
Juan A Imelio

Laboratory of Molecular and Structural Microbiology, Institut Pasteur de Montevideo, Montevideo, Uruguay

Competing interests
The authors declare that no competing interests exist.
Matías R Machado

Biomolecular Simulations, Institut Pasteur de Montevideo, Montevideo, Uruguay

Competing interests
The authors declare that no competing interests exist.
Nicole Larrieux

Laboratory of Molecular and Structural Microbiology, Institut Pasteur de Montevideo, Montevideo, Uruguay

Competing interests
The authors declare that no competing interests exist.
Marcelo A Marti

Dto. Química Biológica e IQUIBICEN-CONICET, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina

Competing interests
The authors declare that no competing interests exist.
Gonzalo Obal

Protein Biophysics Unit, Institut Pasteur de Montevideo, Montevideo, Uruguay

Competing interests
The authors declare that no competing interests exist.
Ariel E Mechaly

Laboratory of Molecular and Structural Microbiology, Institut Pasteur de Montevideo, Montevideo, Uruguay

Competing interests
The authors declare that no competing interests exist.
Alejandro Buschiazzo

Laboratory of Molecular and Structural Microbiology, Institut Pasteur de Montevideo, Montevideo, Uruguay

For correspondence
alebus@pasteur.edu.uy

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-2509-6526

Funding

Agencia Nacional de Investigación e Innovación (FCE2009_1_2679)

Felipe Trajtenberg
Alejandro Buschiazzo

Agence Nationale de la Recherche (PCV06_138918)

Alejandro Buschiazzo

FOCEM (COF 03/11)

Alejandro Buschiazzo

Centro de Biologia Estructural del Mercosur

Alejandro Buschiazzo

Agencia Nacional de Investigación e Innovación (FCE2007_219)

Felipe Trajtenberg
Alejandro Buschiazzo

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.