Molecular association of glucose-6-phosphate isomerase and pyruvate kinase M2 with glyceraldehyde-3-phosphate dehydrogenase in cancer cells
| Autor: | Arup K Bag, Manju Ray, Siddhartha S. Jana, Provas Das, Chitra Mandal, Subhankar Ray, Shekhar Saha, Alok Ghosh, Mahua Rani Das, Sumit K. Dey, Saikat Chakrabarti |
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| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Cancer Research Pyruvate dehydrogenase kinase Pyruvate Kinase Molecular association Gene Expression PKM2 Mass Spectrometry Ehrlich ascites carcinoma Mice 03 medical and health sciences chemistry.chemical_compound Enzyme activator 0302 clinical medicine stomatognathic system Neoplasms Genetics Animals Protein Interaction Domains and Motifs Carcinoma Ehrlich Tumor Glyceraldehyde 3-phosphate dehydrogenase Pyruvate kinase M2 biology Glucose-6-Phosphate Isomerase Glyceraldehyde-3-Phosphate Dehydrogenases Malignancy Pyruvaldehyde Molecular biology Enzyme Activation Disease Models Animal 030104 developmental biology Oncology Biochemistry Glucose 6-phosphate chemistry 030220 oncology & carcinogenesis Cancer cell biology.protein Glyceraldehyde-3-phosphate dehydrogenase Pyruvate kinase Protein Binding Research Article |
| Zdroj: | BMC Cancer |
| ISSN: | 1471-2407 |
| DOI: | 10.1186/s12885-016-2172-x |
| Popis: | Background For a long time cancer cells are known for increased uptake of glucose and its metabolization through glycolysis. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key regulatory enzyme of this pathway and can produce ATP through oxidative level of phosphorylation. Previously, we reported that GAPDH purified from a variety of malignant tissues, but not from normal tissues, was strongly inactivated by a normal metabolite, methylglyoxal (MG). Molecular mechanism behind MG mediated GAPDH inhibition in cancer cells is not well understood. Methods GAPDH was purified from Ehrlich ascites carcinoma (EAC) cells based on its enzymatic activity. GAPDH associated proteins in EAC cells and 3-methylcholanthrene (3MC) induced mouse tumor tissue were detected by mass spectrometry analysis and immunoprecipitation (IP) experiment, respectively. Interacting domains of GAPDH and its associated proteins were assessed by in silico molecular docking analysis. Mechanism of MG mediated GAPDH inactivation in cancer cells was evaluated by measuring enzyme activity, Circular dichroism (CD) spectroscopy, IP and mass spectrometry analyses. Result Here, we report that GAPDH is associated with glucose-6-phosphate isomerase (GPI) and pyruvate kinase M2 (PKM2) in Ehrlich ascites carcinoma (EAC) cells and also in 3-methylcholanthrene (3MC) induced mouse tumor tissue. Molecular docking analyses suggest C-terminal domain preference for the interaction between GAPDH and GPI. However, both C and N termini of PKM2 might be interacting with the C terminal domain of GAPDH. Expression of both PKM2 and GPI is increased in 3MC induced tumor compared with the normal tissue. In presence of 1 mM MG, association of GAPDH with PKM2 or GPI is not perturbed, but the enzymatic activity of GAPDH is reduced to 26.8 ± 5 % in 3MC induced tumor and 57.8 ± 2.3 % in EAC cells. Treatment of MG to purified GAPDH complex leads to glycation at R399 residue of PKM2 only, and changes the secondary structure of the protein complex. Conclusion PKM2 may regulate the enzymatic activity of GAPDH. Increased enzymatic activity of GAPDH in tumor cells may be attributed to its association with PKM2 and GPI. Association of GAPDH with PKM2 and GPI could be a signature for cancer cells. Glycation at R399 of PKM2 and changes in the secondary structure of GAPDH complex could be one of the mechanisms by which GAPDH activity is inhibited in tumor cells by MG. Electronic supplementary material The online version of this article (doi:10.1186/s12885-016-2172-x) contains supplementary material, which is available to authorized users. |
| Databáze: | OpenAIRE |
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