Mechanism of complexing of chlorin e 6 and tetra(p-carboxyphenyl)porphyrin with malate dehydrogenase

Abstract

The mechanism of complexing of the tetrapyrrole-nature photodynamic sensitizers chlorin e6 and tetra(p-carboxyphenyl)porphyrin (TCPP) with the Krebs-cycle enzyme malate dehydrogenase (MDG) has been investigated by spectral-luminescence methods. It is shown that each subunit entering into the composition of the MDG dimer molecule forms an equilibrium complex with one dye molecule. However, if the sites of bonding of TCPP on a protein molecule are independent, the MDG-chlorin e6 complex has a negative cooperativity since after the dye is incorporated into the first subunit of the macromolecule, its penetration to the second subunit becomes difficult. This is explained by the fact that conformation transformations, arising as a result of the incorporation of chlorin into one of the MDG subunits, are transferred to the site of its bonding on other MDG subunits through the intersubunit contacts of the enzyme. It has been established that photosensitizers compete with the hydrophobic fluorescent probe 8-aniline-1-naphthalenesulfonate (ANS) (whose position in the MDG macromolecule was described in detail in the literature) for the sites of bonding on the protein molecule, which allows the conclusion that TCPP and chlorin e6 are localized in the catalytic domain of MDG. In this case, the sites of bonding of these dyes and the sites of bonding of nicotinamide-adenine dinucleotide, occupying the MDG domain that bonds coenzymes, do not interact with each other.