Fig. 1

Characterization of natural porphyrins PpIX and hemin in cells and their physical interaction with G4 structures. A Scheme of natural porphyrins synthesis from δ-aminolevulinic acid (ALA). B,C LC-MS/MS analysis of PpIX. Total ion chromatogram (TIC) of PpIX was generated with precursor m/z 564.2 (B) and the products m/z were 432.28, 490.22, and 505.19 (C). D Standard curve for PpIX LC-MS/MS quantification using peak areas. E, F Mass spectrometry analysis of hemin standards. TIC of hemin was generated with precursor m/z 616 (E) and the products m/z were 483.18, 498.14, and 557.21 (F). G Standard curve for hemin quantification with peak areas using LC-MS/MS analysis. H Workflow for extraction and quantification of PpIX and hemin. Collected cells were disrupted and extracted with acetonitrile for free porphyrins. The pellets containing chromatins and insoluble proteins were further hydrolyzed with acid and extracted with acetonitrile for bound porphyrins. I Quantification of PpIX and hemin in HEK293T, HeLa, HCT116, LM2, U937, and K562 cells. J Characterization of G4-porphyrins interaction with Bio-layer interferometry assays. Biotinylated MYC-G4 was immobilized on streptavidin biosensors and incubated with a range of PpIX or hemin to measure the response in a Gator instrument. K, L Bio-layer interferometry analysis of MYC-G4 with PpIX (K) and hemin (L). The dissociation constant KD values for MYC-G4-PpIX and MYC-G4-hemin were 3.04±0.05 μM and 1.52±0.04 μM, respectively