Carboranyl-Chlorin e6 as a Potent Antimicrobial Photosensitizer

Antimicrobial photodynamic inactivation is currently being widely considered as alternative to antibiotic chemotherapy of infective diseases, attracting much attention to design of novel effective photosensitizers. Carboranyl-chlorin-e₆ (the conjugate of chlorin e₆ with carborane), applied here for the first time for antimicrobial photodynamic inactivation, appeared to be much stronger than chlorin e₆ against Gram-positive bacteria, such as Bacillus subtilis, Staphyllococcus aureus and Mycobacterium sp. Confocal fluorescence spectroscopy and membrane leakage experiments indicated that bacteria cell death upon photodynamic treatment with carboranyl-chlorin-e₆ is caused by loss of cell membrane integrity. The enhanced photobactericidal activity was attributed to the increased accumulation of the conjugate by bacterial cells, as evaluated both by centrifugation and fluorescence correlation spectroscopy. Gram-negative bacteria were rather resistant to antimicrobial photodynamic inactivation mediated by carboranyl-chlorin-e₆. Unlike chlorin e₆, the conjugate showed higher (compared to the wild-type strain) dark toxicity with Escherichia coli ΔtolC mutant, deficient in TolC-requiring multidrug efflux transporters.     

Protection of Saccharomyces cerevisiae against Oxidative and Radiation-Caused Damage by Alkylhydroxybenzenes

The effects of C₇-alkylhydroxybenzene (₇-AHB) and p-hydroxyethylphenol (tyrosol), chemical analogs of microbial anabiosis autoregulators, on the viability of yeast cells under oxidative stress were investigated. The stress was caused by reactive oxygen species (ROS) produced under irradiation of cell suspensions using doses of 10–150 krad at an intensity of 194 rad/s or by singlet oxygen generated in cells photosensitized with chlorin e ₆ (10 g/l). C₇-AHB was found to exert a protective effect. The addition of 0.05–0.16 vol % of C₇-AHB to cell suspensions 30 min before irradiation protected yeast cells from radiation (50 krad). The protective effect of C₇-AHB manifested itself both in the preservation of cell viability during irradiation and in the recovery of their capacity to proliferate after irradiation. In our studies on photodynamic cell inactivation, the fact that the phenolic antioxidant C₇-AHB protects cells from intracellular singlet oxygen was revealed for the first time. The analysis of difference absorption spectra of oxidized derivatives of C₇-AHB demonstrated that the protective mechanism of ₇-AHB involves the scavenging of ROS resulting from oxidative stress. The fact that tyrosol failed to perform a photoprotective function suggests that the antioxidant properties of microbial ₇-AHB are not related to its chaperon functions. The results obtained make an important addition to the spectrum of known antioxidant and antistress effects of phenolic compounds.     

Effect of 2,2'-dipyridyl on accumulation of protoporphyrin IX and its derivatives in yeast mitochondria and plasma membranes

The iron chelator 2,2'-dipyridyl (0.2 mM) more than fourfold increased the concentration of protoporphyrin IX and also of its zinc-containing complex in mitochondria of the yeast Saccharomyces cerevisiae. Protoporphyrin IX and a chlorine derivative of protoporphyrin IX which fluoresces at 670-675 nm were found in isolated plasma membranes of the yeast grown in the presence of 0.2 mM 2,2'-dipyridyl. The accumulation of endogenous porphyrins resulted in intensification of lipid photoperoxidation in mitochondria and plasma membranes and in a dramatically increased sensitivity of the cells to visible light (400-600 nm). The relative contribution of photodestruction of subcellular structures to photoinduced cell inactivation is discussed.     

Photodynamic Damage to Yeast Subcellular Organelles Induced by Elevated Levels of Endogenous Protoporphyrin IX

The 2,2"-dipyridyl-induced accumulation of protoporphyrin IX in Saccharomyces cerevisiae cells was shown to be accompanied by the photoinhibition of cell respiration and the enhancement of the photoinduced permeability of plasma membranes to the fluorescent dye primuline. The visible-light illumination (at 400–600 nm) of the mitochondria and plasma membranes isolated from yeast cells with a high level of endogenous protoporphyrin IX intensified lipid peroxidation in these subcellular organelles. Comparative studies showed that the rad 52 mutant cells, which are deficient in the postreplicative recombinational DNA repair system, are considerably more sensitive to the inactivating action of visible light than are the wild-type cells and the rad 3 mutant cells, which are deficient in the excision DNA repair system. The contribution of photodynamic damage to the yeast subcellular organelles to the lethal photodynamic effect is discussed.     

Light-induced processes of cell protection against photodamage

The processes of light-induced cell protection against photodamage, including DNA photorepair with involvement of photolyases and photoactivation of enzymatic synthesis of photoprotectors and antioxidants (flavonoids, carotenoids, melanins, serotonin, and hemoxygenase), are discussed. Mechanisms of photoprotective effects against photodynamic oxidation of membrane components and photodamage of DNA are analyzed.     

The effect of some antiseptic drugs on the energy transfer in chromatophore photosynthetic membranes of purple non-sulfur bacteria Rhodobacter sphaeroides

Photosynthesis Research - Chromatophores of purple non-sulfur bacteria (PNSB) are invaginations of the cytoplasmic membrane that contain a relatively simple system of light-harvesting...     

Biological photoreceptors of light-dependent regulatory processes

Progress in understanding primary mechanisms of light reception in photoregulatory processes is achieved through discovering new biological photoreceptors, chiefly the regulatory sensors of blue/UV-A light. Among them are LOV domain-containing proteins and DNA photolyase-like cryptochromes, which constitute two widespread groups of photoreceptors that use flavin cofactors (FMN or FAD) as the photoactive chromophores. Bacterial LOV domain modules are connected in photoreceptor proteins with regulatory domains such as diguanylate cyclases/phosphodiesterases, histidine kinases, and DNA-binding domains that are activated by photoconversions of flavin. Identification of red/far-red light sensors in chemotrophic bacteria (bacteriophytochromes) and crystal structures of their photosensor module with bilin chromophore are significant for decoding the mechanisms of phytochrome receptor photoconversion and early step mechanisms of phytochrome-mediated signaling. The only UV-B regulatory photon sensor, UVR8, recently identified in plants, unlike other photoreceptors functions without a prosthetic chromophore: tryptophans of the unique UVR8 protein structure provide a “UV-B antenna”. Our analysis of new data on photosensory properties of the identified photoreceptors in conjunction with their structure opens insight on the influence of the molecular microenvironment on light-induced chromophore reactions, the mechanisms by which the photoactivated chromophores trigger conformational changes in the surrounding protein structure, and structural bases of propagation of these changes to the interacting effector domains/proteins.     

The Photodynamic Inactivation of the Yeast Candida guilliermondii in the Presence of Photodithazine

Photodithazine, a glucosamine salt of chlorin e ₆, is highly effective in sensitization of Candida guilliermondii cells to visible light. The sensitizing effect of photodithazine was found to be related to free or cell surface–bound molecules of this dye. Sodium azide (a singlet oxygen quencher) and propyl gallate (an inhibitor of lipid peroxidation) protected yeast cells from the photodithazine-enhanced photoinactivation.