Time-resolved infra-red studies of photo-excited porphyrins in the presence of nucleic acids and in HeLa tumour cells: insights into binding site and electron transfer dynamics.

Cationic porphyrins based on the 5,10,15,20--(tetrakis-4--methylpyridyl) core (TMPyP4) have been studied extensively over many years due to their strong interactions with a variety of nucleic acid structures, and their potential use as photodynamic therapeutic agents and telomerase inhibitors. In this paper, the interactions of metal-free TMPyP4 and Pt(II)TMPyP4 with guanine-containing nucleic acids are studied for the first time using time-resolved infrared spectroscopy (TRIR). In DO solution (where the metal-free form exists as DTMPyP4) both compounds yielded similar TRIR spectra (between 1450-1750 cm) following pulsed laser excitation in their Soret B-absorption bands. Density functional theory calculations reveal that vibrations centred on the methylpyridinium groups are responsible for the dominant feature at 1640 cm. TRIR spectra of DTMPyP4 or PtTMPyP4 in the presence of guanosine 5'-monophosphate (GMP), double-stranded {d(GC)} or {d(CGCAAATTTGCG)} contain negative-going signals, 'bleaches', indicative of binding close to guanine. TRIR signals for DTMPyP4 or PtTMPyP bound to the quadruplex-forming cMYC sequence {d(TAGGGAGGG)}T indicate that binding occurs on the stacked guanines. For DTMPyP4 bound to guanine-containing systems, the TRIR signal at 1640 cm decays on the picosecond timescale, consistent with electron transfer from guanine to the singlet excited state of DTMPyP4, although IR marker bands for the reduced porphyrin/oxidised guanine were not observed. When PtTMPyP is incorporated into HeLa tumour cells, TRIR studies show protein binding with time-dependent ps/ns changes in the amide absorptions demonstrating TRIR's potential for studying light-activated molecular processes not only with nucleic acids in solution but also in biological cells.