title: Aspirin inhibits glucose‑6‑phosphate dehydrogenase activity in HCT 116 cells through acetylation: Identification of aspirin-acetylated sites.
abstract: Glucose-6-phosphate dehydrogenase (G6PD) catalyzes the first reaction in the pentose phosphate pathway, and generates ribose sugars, which are required for nucleic acid synthesis, and nicotinamide adenine dinucleotide phosphate (NADPH), which is important for neutralization of oxidative stress. The expression of G6PD is elevated in several types of tumor, including colon, breast and lung cancer, and has been implicated in cancer cell growth. Our previous study demonstrated that exposure of HCT 116 human colorectal cancer cells to aspirin caused acetylation of G6PD, and this was associated with a decrease in its enzyme activity. In the present study, this observation was expanded to HT‑29 colorectal cancer cells, in order to compare aspirin‑mediated acetylation of G6PD and its activity between HCT 116 and HT‑29 cells. In addition, the present study aimed to determine the acetylation targets of aspirin on recombinant G6PD to provide an insight into the mechanisms of inhibition. The results demonstrated that the extent of G6PD acetylation was significantly higher in HCT 116 cells compared with in HT‑29 cells; accordingly, a greater reduction in G6PD enzyme activity was observed in the HCT 116 cells. Mass spectrometry analysis of aspirin‑acetylated G6PD (isoform a) revealed that aspirin acetylated a total of 14 lysine residues, which were dispersed throughout the length of the G6PD protein. One of the important amino acid targets of aspirin included lysine 235 (K235, in isoform a) and this corresponds to K205 in isoform b, which has previously been identified as being important for catalysis. Acetylation of G6PD at several sites, including K235 (K205 in isoform b), may mediate inhibition of G6PD activity, which may contribute to the ability of aspirin to exert anticancer effects through decreased synthesis of ribose sugars and NADPH.
authors.affiliation: Department of Pharmaceutical Sciences, South Dakota State University College of Pharmacy, Avera Health and Sciences Center, Brookings, SD 57007, USA.
authors.forename: Guoqiang
authors.initials: G
authors.lastname: Ai
authors.affiliation: Department of Pharmaceutical Sciences, South Dakota State University College of Pharmacy, Avera Health and Sciences Center, Brookings, SD 57007, USA.
authors.forename: Rakesh
authors.initials: R
authors.lastname: Dachineni
authors.affiliation: Department of Pharmaceutical Sciences, South Dakota State University College of Pharmacy, Avera Health and Sciences Center, Brookings, SD 57007, USA.
authors.forename: D Ramesh
authors.initials: DR
authors.lastname: Kumar
authors.affiliation: D'Youville College School of Pharmacy, Buffalo, NY 14201, USA.
authors.forename: Lloyd F
authors.initials: LF
authors.lastname: Alfonso
authors.affiliation: Department of Pharmaceutical Sciences, South Dakota State University College of Pharmacy, Avera Health and Sciences Center, Brookings, SD 57007, USA.
authors.forename: Srinivasan
authors.initials: S
authors.lastname: Marimuthu
authors.affiliation: Department of Pharmaceutical Sciences, South Dakota State University College of Pharmacy, Avera Health and Sciences Center, Brookings, SD 57007, USA.
authors.forename: G Jayarama
authors.initials: GJ
authors.lastname: Bhat
mesh_terms: D000107:Acetylation / Q000187:drug effects; D000596:Amino Acids; D001241:Aspirin / Q000737:chemistry / Q000494:pharmacology*; D001665:Binding Sites; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D005954:Glucosephosphate Dehydrogenase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism*; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D008958:Models, Molecular; D008968:Molecular Conformation; D010427:Pentose Phosphate Pathway / Q000187:drug effects; D011485:Protein Binding
chemical_list: D000596:Amino Acids; D004791:Enzyme Inhibitors; D005954:Glucosephosphate Dehydrogenase; D001241:Aspirin