Extraction and detection of guanosine 5′-diphosphate-3′-diphosphate in amino acid starvation cells of Clavibacter michiganensis
| Autor: | Xing Chen, Laixin Luo, Kaihong Bai, Jianqiang Li, Na Jiang, Qingyang Lyu |
|---|---|
| Rok vydání: | 2021 |
| Předmět: |
Formic acid
Stringent response Guanosine Guanosine Tetraphosphate Microbiology 03 medical and health sciences chemistry.chemical_compound Fungal and Bacterial Physiology - Research Paper Tandem Mass Spectrometry Media Technology heterocyclic compounds Amino Acids 030304 developmental biology chemistry.chemical_classification 0303 health sciences Chromatography biology 030306 microbiology Methanol Extraction (chemistry) Clavibacter biology.organism_classification Amino acid Diphosphates chemistry bacteria Lysozyme Clavibacter michiganensis Chromatography Liquid Alarmone |
| Zdroj: | Braz J Microbiol |
| ISSN: | 1678-4405 1517-8382 |
| DOI: | 10.1007/s42770-021-00488-1 |
| Popis: | Guanosine 5′-diphosphate-3′-diphosphate (ppGpp) is a small molecule nucleotide alarmone that can accumulate under the amino acid starvation state and trigger the stringent response. This study reported the extraction of ppGpp from the Gram-positive bacteria Clavibacter michiganensis through methods using formic acid, lysozyme, or methanol. Following extraction, ppGpp was detected through ultra-high-performance liquid chromatography (UHPLC) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The methanol method showed the highest extraction efficiency for ppGpp among the three methods tested. C. michiganensis cells in exponential growth phase was induced in amino acid starvation by serine hydroxamate (SHX) and used for ppGpp extraction and detection. When using the methanol extraction method, the results showed that ppGpp concentrations in SHX-treated samples were 15.645 nM, 17.656 nM, 20.372 nM, and 19.280 nM at 0 min, 15 min, 30 min and 1 h, respectively, when detected using LC-MS/MS. This is the first report on ppGpp extraction and detection in Clavibacter providing a new idea and approach for nucleotide detection and extraction in bacteria. |
| Databáze: | OpenAIRE |
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