Effects of Visible‐Light Irradiation of Protoporphyrin IX on the Self‐Assembly of Tubulin Heterodimers

Abstract

The formation and the effects of the laser irradiation of the complex formed by protoporphyrin IX (PPIX) and tubulin was investigated. We have used tubulin as a model protein to investigate whether docked photoactive ligands can affect the structure and function of polypeptides upon exposure to visible light. We observed that laser irradiation in the Soret band prompts bleaching of the PPIX which is accompanied by a sharp decrease in the intensity and average fluorescence lifetime of the protein (dominated by the four tryptophan residues of the tubulin monomer). The kinetics indicate non-trivial effects and suggest that the photosensitization of the PPIX bound to tubulin prompts structural alterations of the protein. These modifications were also observed through changes in the energy transfer between Trp residues and PPIX. The result suggest that laser irradiation produces localized partial unfolding of tubulin and that the changes prompt modification of the formation of microtubules in vitro. Measurements of singlet oxygen formation were inconclusive in determining whether the changes are prompted by reactive oxygen species or other excited state mechanisms.

Bacteroides fragilis expresses three proteins similar to Porphyromonas gingivalisHmuY: Hemophore‐like proteins differentially evolved to participate in heme acquisition in oral and gut microbiomes

“…Deuteroheme (FeDPIX) and mesoheme (FeMPIX) (Frontier Specialty Chemicals, Logan, UT, USA) were prepared as described previously 85,86 . PPIX (Sigma‐Aldrich, St. Louis, MO, USA) stock solution was prepared by dissolving PPIX in pure DMSO (99.9%) (Fluka, Munich, Germany) and its concentration was determined in DMSO using the empirical molar absorption coefficient ε 405nm = 150 mM −1 cm −1 87 . CPIII (Frontier Scientific, Logan, UT, USA) stock solution was prepared by dissolving in 0.1 M NaOH and the concentration was determined in 50 mM HEPES pH 7.5, 88 using the empirical molar absorption coefficient ε 393nm = 150.74 mM −1 cm −1 .…”