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- meso -(tetrakis-4- N -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 D 2 O solution (where the metal-free form exists as D 2 TMPyP4) both compounds yielded similar TRIR spectra (between 1450-1750 cm -1 ) 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 ca. 1640 cm -1 . TRIR spectra of D 2 TMPyP4 or PtTMPyP4 in the presence of guanosine 5'-monophosphate (GMP), double-stranded {d(GC) 5 } 2 or {d(CGCAAATTTGCG)} 2 contain negative-going signals, 'bleaches', indicative of binding close to guanine. TRIR signals for D 2 TMPyP4 or PtTMPyP bound to the quadruplex-forming cMYC sequence {d(TAGGGAGGG)} 2 T indicate that binding occurs on the stacked guanines. For D 2 TMPyP4 bound to guanine-containing systems, the TRIR signal at ca. 1640 cm -1 decays on the picosecond timescale, consistent with electron transfer from guanine to the singlet excited state of D 2 TMPyP4, 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.