Affinity-based capture and identification of protein effectors of the growth regulator ppGpp

Boyuan Wang, Peng Dai, David Ding, Amanda Del Rosario, Robert A. Grant, Bradley L. Pentelute, Michael T. Laub

Research output: Contribution to journalArticlepeer-review

84 Scopus citations

Abstract

The nucleotide ppGpp is a highly conserved regulatory molecule in bacteria that helps tune growth rate to nutrient availability. Despite decades of study, how ppGpp regulates growth remains poorly understood. Here, we developed and validated a capture-compound mass spectrometry approach that identified >50 putative ppGpp targets in Escherichia coli. These targets control many key cellular processes and include 13 enzymes required for nucleotide synthesis. We demonstrated that ppGpp inhibits the de novo synthesis of all purine nucleotides by directly targeting the enzyme PurF. By solving a structure of PurF bound to ppGpp, we designed a mutation that ablates ppGpp-based regulation, leading to dysregulation of purine-nucleotide synthesis following ppGpp accumulation. Collectively, our results provide new insights into ppGpp-based growth control and a nearly comprehensive set of targets for future exploration. The capture compounds developed should also enable the rapid identification of ppGpp targets in any species, including pathogens.

Original languageEnglish (US)
Pages (from-to)141-150
Number of pages10
JournalNature chemical biology
Volume15
Issue number2
DOIs
StatePublished - Feb 1 2019
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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