Structural basis for kinase inhibition in the tripartite E. coli HipBST toxin-antitoxin system

René L Bærentsen; Stine V Nielsen; Ragnhild B Skjerning; Jeppe Lyngsø; Francesco Bisiak; Jan Skov Pedersen; Kenn Gerdes; Michael A Sørensen; Ditlev E Brodersen

doi:10.7554/eLife.90400

Structural basis for kinase inhibition in the tripartite E. coli HipBST toxin-antitoxin system

René L Bærentsen, Stine V Nielsen, Ragnhild B Skjerning, Jeppe Lyngsø, Francesco Bisiak, Jan Skov Pedersen, Kenn Gerdes, Michael A Sørensen^*, Ditlev E Brodersen^*

^*Corresponding author for this work

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaper › Journal article › Research › peer-review

6 Citations (Scopus)

Abstract

Many bacteria encode multiple toxin-antitoxin (TA) systems targeting separate, but closely related, cellular functions. The toxin of the Escherichia coli hipBA system, HipA, is a kinase that inhibits translation via phosphorylation of glutamyl-tRNA synthetase. Enteropathogenic E. coli O127:H6 encodes the hipBA-like, tripartite TA system; hipBST, in which the HipT toxin specifically targets the tryptophanyl-tRNA synthetase, TrpS. Notably, in the tripartite system, the function as antitoxin has been taken over by the third protein, HipS, but the molecular details of how activity of HipT is inhibited remain poorly understood. Here, we show that HipBST is structurally different from E. coli HipBA and that the unique HipS protein, which is homologous to the N-terminal subdomain of HipA, inhibits the kinase through insertion of a conserved Trp residue into the active site. We also show how auto-phosphorylation at two conserved sites in the kinase toxin serve different roles and affect the ability of HipS to neutralize HipT. Finally, solution structural studies show how phosphorylation affects overall TA complex flexibility.

Original language	English
Article number	RP90400
Journal	eLife
Volume	12
Number of pages	20
ISSN	2050-084X
DOIs	https://doi.org/10.7554/eLife.90400
Publication status	Published - 6 Nov 2023

Keywords

Antitoxins/metabolism
Escherichia coli Proteins/metabolism
Escherichia coli/genetics
Phosphorylation
Toxin-Antitoxin Systems/genetics

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10.7554/eLife.90400Licence: CC BY

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title = "Structural basis for kinase inhibition in the tripartite E. coli HipBST toxin-antitoxin system",

abstract = "Many bacteria encode multiple toxin-antitoxin (TA) systems targeting separate, but closely related, cellular functions. The toxin of the Escherichia coli hipBA system, HipA, is a kinase that inhibits translation via phosphorylation of glutamyl-tRNA synthetase. Enteropathogenic E. coli O127:H6 encodes the hipBA-like, tripartite TA system; hipBST, in which the HipT toxin specifically targets the tryptophanyl-tRNA synthetase, TrpS. Notably, in the tripartite system, the function as antitoxin has been taken over by the third protein, HipS, but the molecular details of how activity of HipT is inhibited remain poorly understood. Here, we show that HipBST is structurally different from E. coli HipBA and that the unique HipS protein, which is homologous to the N-terminal subdomain of HipA, inhibits the kinase through insertion of a conserved Trp residue into the active site. We also show how auto-phosphorylation at two conserved sites in the kinase toxin serve different roles and affect the ability of HipS to neutralize HipT. Finally, solution structural studies show how phosphorylation affects overall TA complex flexibility.",

keywords = "Antitoxins/metabolism, Escherichia coli Proteins/metabolism, Escherichia coli/genetics, Phosphorylation, Toxin-Antitoxin Systems/genetics",

author = "B{\ae}rentsen, {Ren{\'e} L} and Nielsen, {Stine V} and Skjerning, {Ragnhild B} and Jeppe Lyngs{\o} and Francesco Bisiak and Pedersen, {Jan Skov} and Kenn Gerdes and S{\o}rensen, {Michael A} and Brodersen, {Ditlev E}",

year = "2023",

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T1 - Structural basis for kinase inhibition in the tripartite E. coli HipBST toxin-antitoxin system

AU - Bærentsen, René L

AU - Nielsen, Stine V

AU - Skjerning, Ragnhild B

AU - Lyngsø, Jeppe

AU - Bisiak, Francesco

AU - Pedersen, Jan Skov

AU - Gerdes, Kenn

AU - Sørensen, Michael A

AU - Brodersen, Ditlev E

PY - 2023/11/6

Y1 - 2023/11/6

N2 - Many bacteria encode multiple toxin-antitoxin (TA) systems targeting separate, but closely related, cellular functions. The toxin of the Escherichia coli hipBA system, HipA, is a kinase that inhibits translation via phosphorylation of glutamyl-tRNA synthetase. Enteropathogenic E. coli O127:H6 encodes the hipBA-like, tripartite TA system; hipBST, in which the HipT toxin specifically targets the tryptophanyl-tRNA synthetase, TrpS. Notably, in the tripartite system, the function as antitoxin has been taken over by the third protein, HipS, but the molecular details of how activity of HipT is inhibited remain poorly understood. Here, we show that HipBST is structurally different from E. coli HipBA and that the unique HipS protein, which is homologous to the N-terminal subdomain of HipA, inhibits the kinase through insertion of a conserved Trp residue into the active site. We also show how auto-phosphorylation at two conserved sites in the kinase toxin serve different roles and affect the ability of HipS to neutralize HipT. Finally, solution structural studies show how phosphorylation affects overall TA complex flexibility.

AB - Many bacteria encode multiple toxin-antitoxin (TA) systems targeting separate, but closely related, cellular functions. The toxin of the Escherichia coli hipBA system, HipA, is a kinase that inhibits translation via phosphorylation of glutamyl-tRNA synthetase. Enteropathogenic E. coli O127:H6 encodes the hipBA-like, tripartite TA system; hipBST, in which the HipT toxin specifically targets the tryptophanyl-tRNA synthetase, TrpS. Notably, in the tripartite system, the function as antitoxin has been taken over by the third protein, HipS, but the molecular details of how activity of HipT is inhibited remain poorly understood. Here, we show that HipBST is structurally different from E. coli HipBA and that the unique HipS protein, which is homologous to the N-terminal subdomain of HipA, inhibits the kinase through insertion of a conserved Trp residue into the active site. We also show how auto-phosphorylation at two conserved sites in the kinase toxin serve different roles and affect the ability of HipS to neutralize HipT. Finally, solution structural studies show how phosphorylation affects overall TA complex flexibility.

KW - Antitoxins/metabolism

KW - Escherichia coli Proteins/metabolism

KW - Escherichia coli/genetics

KW - Phosphorylation

KW - Toxin-Antitoxin Systems/genetics

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U2 - 10.7554/eLife.90400

DO - 10.7554/eLife.90400

M3 - Journal article

C2 - 37929938

SN - 2050-084X

VL - 12

JO - eLife

JF - eLife

M1 - RP90400

ER -

Structural basis for kinase inhibition in the tripartite E. coli HipBST toxin-antitoxin system

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