New potential eukaryotic substrates of the mycobacterial protein tyrosine phosphatase PtpA: hints of a bacterial modulation of macrophage bioenergetics state
| Autor: | María Magdalena Portela, Gonzalo Obal, Anne-Marie Labandera, Ana M. Ferreira, Federico Carrion, Magdalena Gil, Mariana Margenat, Hernán Terenzi, Andrea Villarino, Guilherme Razzera, Otto Pritsch, Rosario Durán, Marcela Purificação |
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| Jazyk: | angličtina |
| Rok vydání: | 2015 |
| Předmět: |
Bioenergetics
education Plasma protein binding Protein tyrosine phosphatase Biology medicine.disease_cause Article Mass Spectrometry law.invention Cell Line Substrate Specificity Bacterial Proteins law Protein Interaction Mapping medicine Macrophage Humans Phosphorylation Mutation Multidisciplinary Circular Dichroism Macrophages female genital diseases and pregnancy complications Recombinant Proteins Cell biology Biochemistry Cell culture Recombinant DNA Protein Tyrosine Phosphatases Protein Binding |
| Zdroj: | Scientific Reports |
| ISSN: | 2045-2322 |
| Popis: | The bacterial protein tyrosine phosphatase PtpA is a key virulence factor released by Mycobacterium tuberculosis in the cytosol of infected macrophages. So far only two unrelated macrophage components (VPS33B, GSK3α) have been identified as PtpA substrates. As tyrosine phosphatases are capable of using multiple substrates, we developed an improved methodology to pull down novel PtpA substrates from an enriched P-Y macrophage extract using the mutant PtpA D126A. This methodology reduced non-specific protein interactions allowing the identification of four novel putative PtpA substrates by MALDI-TOF-MS and nano LC-MS: three mitochondrial proteins - the trifunctional enzyme (TFP), the ATP synthase and the sulfide quinone oxidoreductase - and the cytosolic 6-phosphofructokinase. All these proteins play a relevant role in cell energy metabolism. Using surface plasmon resonance, PtpA was found to bind immunopurified human TFP through its catalytic site since TFP-PtpA association was inhibited by a specific phosphatase inhibitor. Moreover, PtpA wt was capable of dephosphorylating immunopurified human TFP in vitro supporting that TFP may be a bona fide PtpA susbtrate. Overall, these results suggest a novel scenario where PtpA-mediated dephosphorylation may affect pathways involved in cell energy metabolism, particularly the beta oxidation of fatty acids through modulation of TFP activity and/or cell distribution. |
| Databáze: | OpenAIRE |
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