(p)ppGpp controls stringent factors by exploiting antagonistic allosteric coupling between catalytic domains

Mohammad Roghanian; Katleen Van Nerom; Hiraku Takada; Julien Caballero-Montes; Hedvig Tamman; Pavel Kudrin; Ariel Talavera; Ievgen Dzhygyr; Simon Ekström; Gemma C Atkinson; Abel Garcia-Pino; Vasili Hauryliuk

doi:10.1016/j.molcel.2021.07.026

(p)ppGpp controls stringent factors by exploiting antagonistic allosteric coupling between catalytic domains

Mol Cell. 2021 Aug 19;81(16):3310-3322.e6. doi: 10.1016/j.molcel.2021.07.026.

Affiliations

¹ Department of Molecular Biology, Umeå University, 901 87, Umeå, Sweden; Laboratory for Molecular Infection Medicine Sweden, Umeå University, 901 87, Umeå, Sweden. Electronic address: mohammad.roghanian@regionh.dk.
² Cellular and Molecular Microbiology, Faculté des Sciences, Université libre de Bruxelles, Campus La Plaine, Boulevard du Triomphe, 1050, Brussels, Belgium.
³ Department of Molecular Biology, Umeå University, 901 87, Umeå, Sweden; Laboratory for Molecular Infection Medicine Sweden, Umeå University, 901 87, Umeå, Sweden.
⁴ University of Tartu, Institute of Technology, 50411 Tartu, Estonia.
⁵ Department of Biomedical Engineering, Lund University, 221 00 Lund, Sweden.
⁶ Department of Molecular Biology, Umeå University, 901 87, Umeå, Sweden; Department of Experimental Medical Science, Lund University, 221 00 Lund, Sweden.
⁷ Cellular and Molecular Microbiology, Faculté des Sciences, Université libre de Bruxelles, Campus La Plaine, Boulevard du Triomphe, 1050, Brussels, Belgium; WELBIO, Avenue Hippocrate 75, 1200 Brussels, Belgium. Electronic address: abel.garcia.pino@ulb.be.
⁸ Department of Molecular Biology, Umeå University, 901 87, Umeå, Sweden; Laboratory for Molecular Infection Medicine Sweden, Umeå University, 901 87, Umeå, Sweden; University of Tartu, Institute of Technology, 50411 Tartu, Estonia; Department of Experimental Medical Science, Lund University, 221 00 Lund, Sweden. Electronic address: vasili.hauryliuk@med.lu.se.

PMID: 34416138
DOI: 10.1016/j.molcel.2021.07.026

Abstract

Amino acid starvation is sensed by Escherichia coli RelA and Bacillus subtilis Rel through monitoring the aminoacylation status of ribosomal A-site tRNA. These enzymes are positively regulated by their product-the alarmone nucleotide (p)ppGpp-through an unknown mechanism. The (p)ppGpp-synthetic activity of Rel/RelA is controlled via auto-inhibition by the hydrolase/pseudo-hydrolase (HD/pseudo-HD) domain within the enzymatic N-terminal domain region (NTD). We localize the allosteric pppGpp site to the interface between the SYNTH and pseudo-HD/HD domains, with the alarmone stimulating Rel/RelA by exploiting intra-NTD autoinhibition dynamics. We show that without stimulation by pppGpp, starved ribosomes cannot efficiently activate Rel/RelA. Compromised activation by pppGpp ablates Rel/RelA function in vivo, suggesting that regulation by the second messenger (p)ppGpp is necessary for mounting an acute starvation response via coordinated enzymatic activity of individual Rel/RelA molecules. Control by (p)ppGpp is lacking in the E. coli (p)ppGpp synthetase SpoT, thus explaining its weak synthetase activity.

Keywords: (p)ppGpp; Hydrogen-Deuterium exchange Mass Spectrometry; Rel; RelA; RelA-SpoT Homolog; SpoT; allostery; positive feedback regulation; ribosome; stringent response.

Publication types

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

MeSH terms

Allosteric Regulation / genetics*
Amino Acids / metabolism
Bacillus subtilis / genetics
Bacillus subtilis / metabolism
Catalytic Domain / genetics
Escherichia coli / genetics
Escherichia coli / metabolism
Escherichia coli Proteins / genetics*
GTP Pyrophosphokinase / genetics*
Guanosine Pentaphosphate / genetics*
Hydrolases / genetics
Pyrophosphatases / genetics*
Ribosomes / genetics
Ribosomes / metabolism
Starvation / genetics
Starvation / metabolism

Substances

Amino Acids
Escherichia coli Proteins
Guanosine Pentaphosphate
GTP Pyrophosphokinase
relA protein, E coli
Hydrolases
guanosine-3',5'-bis(diphosphate) 3'-pyrophosphatase
Pyrophosphatases