Fluorescent proteins as singlet oxygen photosensitizers: Mechanistic studies in photodynamic inactivation of bacteria

Rubén Ruiz-González, John H. White, Aitziber L. Cortajarena, Montserrat Agut, Santi Nonell, Cristina Flors

Research output: Book chapterConference contributionpeer-review

5 Citations (Scopus)

Abstract

Antimicrobial photodynamic therapy (aPDT) combines a photosensitizer, light and oxygen to produce reactive oxygen species (ROS), mainly singlet oxygen (1O2), to photo-oxidize important biomolecules and induce cell death. aPDT is a promising alternative to standard antimicrobial strategies, but its mechanisms of action are not well understood. One of the reasons for that is the lack of control of the photosensitizing drugs location. Here we report the use of geneticallyencoded fluorescent proteins that are also 1O2 photosensitizers to address the latter issue. First, we have chosen the red fluorescent protein TagRFP as a photosensitizer, which unlike other fluorescent proteins such as KillerRed, is able to produce 1O2 but not other ROS. TagRFP photosensitizes 1O2 with a small, but not negligible, quantum yield. In addition, we have used miniSOG, a more efficient 1O2 photosensitizing fluorescent flavoprotein that has been recently engineered from phototropin 2. We have genetically incorporated these two photosensitizers into the cytosol of E. coli and demonstrated that intracellular 1O 2 is sufficient to kill bacteria. Additional assays have provided further insight into the mechanism of cell death. Photodamage seems to occur primarily in the inner membrane, and extends to the outer membrane if the photosensitizer's efficiency is high enough. These observations are markedly different to those reported for external photosensitizers, suggesting that the site where 1O2 is primarily generated proves crucial for inflicting different types of cell damage.

Original languageEnglish
Title of host publicationReporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications V
DOIs
Publication statusPublished - 2013
EventReporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications V - San Francisco, CA, United States
Duration: 4 Feb 20136 Feb 2013

Publication series

NameProgress in Biomedical Optics and Imaging - Proceedings of SPIE
Volume8596
ISSN (Print)1605-7422

Conference

ConferenceReporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications V
Country/TerritoryUnited States
CitySan Francisco, CA
Period4/02/136/02/13

Keywords

  • TagRFP
  • antimicrobial photodynamic therapy
  • fluorescent proteins
  • miniSOG
  • photosensitization
  • singlet oxygen

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