Nonhydrolysable Analogues of (p)ppGpp and (p)ppApp Alarmone Nucleotides as Novel Molecular Tools

0545506 - ÚOCHB 2022 RIV US eng J - Journal Article
Mojr, Viktor - Roghanian, M. - Tamman, H. - Do Pham, Duy Dinh - Petrová, Magdalena - Pohl, Radek - Takada, H. - Van Nerom, K. - Ainelo, H. - Caballero-Montes, J. - Jimmy, S. - Garcia-Pino, A. - Hauryliuk, V. - Rejman, Dominik
Nonhydrolysable Analogues of (p)ppGpp and (p)ppApp Alarmone Nucleotides as Novel Molecular Tools.
ACS Chemical Biology. Roč. 16, č. 9 (2021), s. 1680-1691. ISSN 1554-8929. E-ISSN 1554-8937
R&D Projects: GA MŠMT(CZ) 8F19006
Institutional support: RVO:61388963
Keywords : pppGpp * pppApp * nonhydrolysable * molecular tools
OECD category: Organic chemistry
Impact factor: 4.634, year: 2021
Method of publishing: Limited access
https://doi.org/10.1021/acschembio.1c00398

While alarmone nucleotides guanosine-3′,5′-bisdiphosphate (ppGpp) and guanosine-5′-triphosphate-3′-diphosphate (pppGpp) are archetypical bacterial second messengers, their adenosine analogues ppApp (adenosine-3′,5′-bisdiphosphate) and pppApp (adenosine-5′-triphosphate-3′-diphosphate) are toxic effectors that abrogate bacterial growth. The alarmones are both synthesized and degraded by the members of the RelA-SpoT Homologue (RSH) enzyme family. Because of the chemical and enzymatic liability of (p)ppGpp and (p)ppApp, these alarmones are prone to degradation during structural biology experiments. To overcome this limitation, we have established an efficient and straightforward procedure for synthesizing nonhydrolysable (p)ppNuNpp analogues starting from 3′-azido-3′-deoxyribonucleotides as key intermediates. To demonstrate the utility of (p)ppGNpp as a molecular tool, we show that (i) as an HD substrate mimic, ppGNpp competes with ppGpp to inhibit the enzymatic activity of human MESH1 Small Alarmone Hyrolase, SAH, and (ii) mimicking the allosteric effects of (p)ppGpp, (p)ppGNpp acts as a positive regulator of the synthetase activity of long ribosome-associated RSHs Rel and RelA. Finally, by solving the structure of the N-terminal domain region (NTD) of T. thermophilus Rel complexed with pppGNpp, we show that as an HD substrate mimic, the analogue serves as a bona fide orthosteric regulator that promotes the same intra-NTD structural rearrangements as the native substrate.
Permanent Link: http://hdl.handle.net/11104/0322183