Format

Send to

Choose Destination
Nat Chem Biol. 2019 Feb;15(2):141-150. doi: 10.1038/s41589-018-0183-4. Epub 2018 Dec 17.

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

Author information

1
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA.
2
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, MA, USA.
3
Koch Institute for Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA.
4
Center for Environmental Health Sciences MIT, Massachusetts Institute of Technology, Cambridge, MA, USA.
5
Broad Institute of Harvard and MIT, Cambridge, MA, USA.
6
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, USA. laub@mit.edu.
7
Howard Hughes Medical Institute, Massachusetts Institute of Technology, Cambridge, MA, USA. laub@mit.edu.

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.

PMID:
30559427
DOI:
10.1038/s41589-018-0183-4

Supplemental Content

Full text links

Icon for Nature Publishing Group
Loading ...
Support Center