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International Journal of Molecular... Dec 2022is a dangerous hospital pathogen primarily due to its ability to form biofilms on different abiotic and biotic surfaces. The present study investigated the effect of...
is a dangerous hospital pathogen primarily due to its ability to form biofilms on different abiotic and biotic surfaces. The present study investigated the effect of riboflavin- and chlorophyllin-based antimicrobial photodynamic therapy, performed with near-ultraviolet or blue light on the viability of bacterial cells in biofilms and their structural stability, also determining the extent of photoinduced generation of intracellular reactive oxygen species as well as the ability of to form biofilms after the treatment. The efficacy of antimicrobial photodynamic therapy was compared with that of light alone and the role of the photosensitizer type on the photosensitization mechanism was demonstrated. We found that the antibacterial effect of riboflavin-based antimicrobial photodynamic therapy depends on the ability of photoactivated riboflavin to generate intracellular reactive oxygen species but does not depend on the concentration of riboflavin and pre-incubation time before irradiation. Moreover, our results suggest a clear interconnection between the inactivation efficiency of chlorophyllin-based antimicrobial photodynamic therapy and the sensitivity of biofilms to used light. In summary, all the analyzed results suggest that riboflavin-based antimicrobial photodynamic therapy and chlorophyllin-based antimicrobial photodynamic therapy have the potential to be applied as an antibacterial treatment against biofilms or as a preventive measure against biofilm formation.
Topics: Acinetobacter baumannii; Reactive Oxygen Species; Anti-Infective Agents; Anti-Bacterial Agents; Photochemotherapy; Biofilms; Riboflavin
PubMed: 36614160
DOI: 10.3390/ijms24010722 -
The FEBS Journal Aug 2011Riboflavin (vitamin B(2)) serves as the precursor for FMN and FAD in almost all organisms that utilize the redox-active isoalloxazine ring system as a coenzyme in... (Review)
Review
Riboflavin (vitamin B(2)) serves as the precursor for FMN and FAD in almost all organisms that utilize the redox-active isoalloxazine ring system as a coenzyme in enzymatic reactions. The role of flavin, however, is not limited to redox processes, as ∼ 10% of flavin-dependent enzymes catalyze nonredox reactions. Moreover, the flavin cofactor is also widely used as a signaling and sensing molecule in biological processes such as phototropism and nitrogen fixation. Here, we present a study of 374 flavin-dependent proteins analyzed with regard to their function, structure and distribution among 22 archaeal, eubacterial, protozoan and eukaryotic genomes. More than 90% of flavin-dependent enzymes are oxidoreductases, and the remaining enzymes are classified as transferases (4.3%), lyases (2.9%), isomerases (1.4%) and ligases (0.4%). The majority of enzymes utilize FAD (75%) rather than FMN (25%), and bind the cofactor noncovalently (90%). High-resolution structures are available for about half of the flavoproteins. FAD-containing proteins predominantly bind the cofactor in a Rossmann fold (∼ 50%), whereas FMN-containing proteins preferably adopt a (βα)(8)-(TIM)-barrel-like or flavodoxin-like fold. The number of genes encoding flavin-dependent proteins varies greatly in the genomes analyzed, and covers a range from ∼ 0.1% to 3.5% of the predicted genes. It appears that some species depend heavily on flavin-dependent oxidoreductases for degradation or biosynthesis, whereas others have minimized their flavoprotein arsenal. An understanding of 'flavin-intensive' lifestyles, such as in the human pathogen Mycobacterium tuberculosis, may result in valuable new intervention strategies that target either riboflavin biosynthesis or uptake.
Topics: Eukaryota; Flavin Mononucleotide; Flavin-Adenine Dinucleotide; Flavins; Flavoproteins; Genomics; Prokaryotic Cells; Riboflavin
PubMed: 21635694
DOI: 10.1111/j.1742-4658.2011.08202.x -
Transfusion and Apheresis Science :... Aug 2022Test the ability of Mirasol Pathogen Reduction Technology (PRT, Terumo BCT, Lakewood Co, USA) treatment with riboflavin and ultraviolet light (R + UV) in reducing...
BACKGROUND
Test the ability of Mirasol Pathogen Reduction Technology (PRT, Terumo BCT, Lakewood Co, USA) treatment with riboflavin and ultraviolet light (R + UV) in reducing SARS-CoV-2 infectivity while maintaining blood product quality.
MATERIAL AND METHODS
SARS-CoV-2 strains were isolated and titrated to prepare cell free virus for plasma units infection. The units were then under treatment with Mirasol PRT. The infectious titers were determined before and after treatment with an in house microtitration assay on Vero E6 cells. Thirty-six plasma pool bags underwent PRT treatment.
RESULTS
In all the experiments, the measured titer following riboflavin and UV treatment was below the limit of detection of microtitration assay for all the different SARS-CoV-2 strains. Despite the high copies number detected by RT-PCR for each viral strain after treatment, viruses were completely inactivated and not able to infect VERO E6 cells.
CONCLUSION
Riboflavin and UV light treatment effectively reduced the virus titers of human plasma to the limit of detection in tissue culture, regardless of the strain. These data suggest that pathogen reduction in blood products highlight the safety of CP therapy procedures for critically ill COVID-19 patients, while maintaining blood product quality.
Topics: Humans; Riboflavin; SARS-CoV-2; Ultraviolet Rays; COVID-19 Drug Treatment
PubMed: 35227599
DOI: 10.1016/j.transci.2022.103398 -
Yonsei Medical Journal 1968
Topics: Humans; Intestinal Absorption; Male; Riboflavin; Thiamine
PubMed: 5715896
DOI: 10.3349/ymj.1968.9.1.11 -
PloS One 2023To investigate the effects of scleral collagen cross-linking (SXL) using riboflavin and ultraviolet A (UVA) light on the scleral thickness of different regions and...
OBJECTIVE
To investigate the effects of scleral collagen cross-linking (SXL) using riboflavin and ultraviolet A (UVA) light on the scleral thickness of different regions and expression of matrix metalloproteinase 2 (MMP-2) and membrane-type MMP-1 (MT1-MMP) in guinea pigs with lens-induced myopia.
METHODS
Forty-eight 4-week-old guinea pigs were assigned to three groups (n = 16 per group): SXL group, lens-induced myopia (LIM) group, and control group. The sclera of the right eye of the guinea pig in the SXL group was surgically exposed, riboflavin was dropped on the treatment area for 10 minutes before the 30-minute UVA irradiation. The same surgical procedure was performed in the LIM group without UVA irradiation. The -10.00 D lenses were then placed on the right eyes of guinea pigs in the SXL and LIM groups for six weeks. The control group received no treatment. The left eyes were untreated in all groups. The ocular axial length (AXL) and refraction were measured at 4 weeks and 10 weeks of age. 10-week-old guinea pigs were sacrificed, and the right eyes were enucleated and evenly divided for preparation of hematoxylin and eosin (HE) stained sections, quantitative real-time polymerase chain reaction (qPCR) and western blotting. The scleral thickness of different regions was measured on HE stained sections. The temporal half of the sclera was harvested to measure the expression of MMP-2 and MT1-MMP by qPCR and western blotting.
RESULTS
The AXL was significantly shorter, and the degree of myopic refraction was significantly lower in the SXL group than those in the LIM group at 10 weeks of age. The scleral thickness of the cross-linked area was significantly greater in the SXL group than that of the corresponding area in the LIM group, while the scleral thickness of the untreated nasal side was not significantly different between the SXL group and the LIM group. The expression of MMP-2 and MT1-MMP of the cross-linked sclera was significantly downregulated compared with that of the corresponding area in the LIM group.
CONCLUSION
Riboflavin/UVA SXL could slow myopia progression and thicken the cross-linked sclera in guinea pigs, which might be related to the downregulation of MMP-2 and MT1-MMP expression during the scleral remodeling process.
Topics: Guinea Pigs; Animals; Sclera; Matrix Metalloproteinase 2; Matrix Metalloproteinase 14; Myopia; Riboflavin; Collagen; Disease Models, Animal
PubMed: 36652495
DOI: 10.1371/journal.pone.0279111 -
Acta Ophthalmologica Jun 2022To determine corneal cross-linking (CXL) efficacy and chromophore penetration after excimer laser-assisted patterned de-epithelialization.
PURPOSE
To determine corneal cross-linking (CXL) efficacy and chromophore penetration after excimer laser-assisted patterned de-epithelialization.
METHODS
Two-hundred-twenty porcine eyes were de-epithelialized ex vivo, either fully (mechanical; n = 88) or patterned (excimer laser; n = 132). Consecutively, corneas were impregnated with hypo- or hyperosmolar riboflavin (RF; n = 20, RF-D; n = 40, respectively) or water-soluble taurine (WST11; n = 40, and WST-D; n = 40, respectively), or kept unimpregnated (n = 80). Sixty corneas were subsequently irradiated, inducing CXL, with paired contralateral eyes serving as controls. Outcome measurements included strip extensiometry to assess CXL efficacy, and spectrophotometry and fluorescence microscopy to determine stromal chromophore penetration.
RESULTS
All tested chromophores induced significant CXL (p < 0.001), ranging from 7.6% to 14.6%, with similar stiffening for all formulations (p = 0.60) and both de-epithelialization methods (p = 0.56). Light transmittance was significantly lower (p < 0.001) after full compared with patterned de-epithelialization. Stromal chromophore penetration was comparable between fully and patterned de-epithelialized samples, with full penetration in RD and RF-D samples and penetration depths measuring 591.7 ± 42.8 µm and 592.9 ± 63.5 µm for WST11 (p = 0.963) and 504.2 ± 43.2 µm and 488.8 ± 93.1 µm for WST-D (p = 0.669), respectively.
CONCLUSIONS
Excimer laser-assisted patterned de-epithelialization allows for effective CXL. Stromal chromophore concentration is, however, reduced, which may have safety implications given the need for sufficient UVA attenuation in RF/UVA CXL. The different safety profile of near-infrared (NIR) may allow safe WST11/NIR CXL even with reduced stromal chromophore concentration values. In vivo studies are needed to evaluate the benefits and further assess safety of excimer laser-assisted patterned de-epithelialization for corneal CXL.
Topics: Animals; Collagen; Cornea; Corneal Stroma; Cross-Linking Reagents; Humans; Lasers, Excimer; Photosensitizing Agents; Riboflavin; Swine; Ultraviolet Rays
PubMed: 34533277
DOI: 10.1111/aos.15021 -
ACS Applied Bio Materials May 2023Bioengineered corneal tissue is a promising therapeutic modality for the treatment of corneal blindness as a substitute for cadaveric graft tissue. In this study, we...
Bioengineered corneal tissue is a promising therapeutic modality for the treatment of corneal blindness as a substitute for cadaveric graft tissue. In this study, we fabricated a collagen gel using ultraviolet-A (UV-A) light and riboflavin as a photosensitizer (PhotoCol-RB) as an -forming matrix to fill corneal wounds and create a cohesive interface between the crosslinked gel and adjacent collagen. The PhotoCol-RB gels supported corneal epithelialization and exhibited higher transparency compared to physically crosslinked collagen. We showed that different riboflavin concentrations yielded gels with different mechanical and biological properties. experiments using human corneal epithelial cells (hCECs) showed that hCECs are able to proliferate on the gel and express corneal cell markers such as cytokeratin 12 (CK12) and tight junctions (ZO-1). Using an burst assay, we also showed that the PhotoCol-RB gels are able to seal corneal perforations. organ culture of the gels filling lamellar keratectomy wounds showed that the epithelium that regenerated over the PhotoCol-RB gels formed a multilayer compared to just a double layer for those that grew over physically cross-linked collagen. These gels can be formed either directly on the wound site to conform to the geometry of a defect, or can be preformed and then applied to the corneal wound. Our results indicate that PhotoCol-RB gels merit further investigation as a way to stabilize and repair deep and perforating corneal wounds.
Topics: Humans; Collagen; Cornea; Regeneration; Riboflavin; Gels
PubMed: 37126648
DOI: 10.1021/acsabm.3c00015 -
Photochemistry and Photobiology 2014Within the last decade new technologies have been developed and implemented which employ light, often in the presence of a photosensitizer, to inactivate pathogens that... (Review)
Review
Within the last decade new technologies have been developed and implemented which employ light, often in the presence of a photosensitizer, to inactivate pathogens that reside in human blood products for the purpose of transfusion. These pathogen reduction technologies attempt to find the proper balance between pathogen kill and cell quality. Each system utilizes various chemistries that not only impact which pathogens they can inactivate and how, but also how the treatments affect the plasma and cellular proteins and to what degree. This paper aims to present the various chemical mechanisms for pathogen reduction in transfusion medicine that are currently practiced or in development.
Topics: Bacteria; Blood Transfusion; Furocoumarins; Humans; Light; Methylene Blue; Photopheresis; Photosensitizing Agents; Riboflavin; Trypanosoma; Viruses
PubMed: 25041351
DOI: 10.1111/php.12311 -
The Plant Journal : For Cell and... Dec 2016Despite the importance of riboflavin as the direct precursor of the cofactors flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN), the physiologically...
Despite the importance of riboflavin as the direct precursor of the cofactors flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN), the physiologically relevant catalyst dephosphorylating the riboflavin biosynthesis pathway intermediate 5-amino-6-ribitylamino-2,4(1H,3H) pyrimidinedione 5'-phosphate (ARPP) has not been characterized from any organism. By using as the query sequence a previously identified plastidial FMN hydrolase AtcpFHy1 (At1g79790), belonging to the haloacid dehalogenase (HAD) superfamily, seven candidates for the missing ARPP phosphatase were found, cloned, recombinantly expressed, and purified. Activity screening showed that the enzymes encoded by AtcpFHy1, At4g11570, and At4g25840 catalyze dephosphorylation of ARPP. AtcpFHy1 was renamed AtcpFHy/PyrP1, At4g11570 and At4g25840 were named AtPyrP2 and AtGpp1/PyrP3, respectively. Subcellular localization in planta indicated that AtPyrP2 was localized in plastids and AtGpp1/PyrP3 in mitochondria. Biochemical characterization of AtcpFHy/PyrP1 and AtPyrP2 showed that they have similar K values for the substrate ARPP, with AtcpFHy/PyrP1 having higher catalytic efficiency. Screening of 21 phosphorylated substrates showed that AtPyrP2 is specific for ARPP. Molecular weights of AtcpFHy/PyrP1 and AtPyrP2 were estimated at 46 and 72 kDa, suggesting dimers. pH and temperature optima for AtcpFHy/PyrP1 and AtPyrP2 were ~7.0-8.5 and 40-50°C. T-DNA knockout of AtcpFHy/PyrP1 did not affect the flavin profile of the transgenic plants, whereas silencing of AtPyrP2 decreased accumulation of riboflavin, FMN, and FAD. Our results strongly support AtPyrP2 as the missing phosphatase on the riboflavin biosynthesis pathway in Arabidopsis thaliana. The identification of this enzyme closes a long-standing gap in understanding of the riboflavin biosynthesis in plants.
Topics: Arabidopsis; Dinitrocresols; Flavin Mononucleotide; Flavin-Adenine Dinucleotide; Hydrolases; Riboflavin; Uracil Nucleotides
PubMed: 27490826
DOI: 10.1111/tpj.13291 -
Journal of the American Chemical Society Aug 2022In proteins, the amino acids Phe, Tyr, and especially Trp are frequently involved in π interactions such as π-π, cation-π, and CH-π bonds. These interactions are...
In proteins, the amino acids Phe, Tyr, and especially Trp are frequently involved in π interactions such as π-π, cation-π, and CH-π bonds. These interactions are often crucial for protein structure and protein-ligand binding. A powerful means to study these interactions is progressive fluorination of these aromatic residues to modulate the electrostatic component of the interaction. However, to date no protein expression platform is available to produce milligram amounts of proteins labeled with such fluorinated amino acids. Here, we present a Trp auxotroph-based expression system for efficient incorporation (≥95%) of mono-, di-, tri-, and tetrafluorinated, as well as a methylated Trp analog. As a model protein we have chosen LmrR, a dimeric multidrug transcriptional repressor protein from LmrR binds aromatic drugs, like daunomycin and riboflavin, between Trp96 and Trp96' in the dimer interface. Progressive fluorination of Trp96 decreased the affinity for the drugs 6- to 70-fold, clearly establishing the importance of electrostatic π-π interactions for drug binding. Presteady state kinetic data of the LmrR-drug interaction support the enthalpic nature of the interaction, while high resolution crystal structures of the labeled protein-drug complexes provide for the first time a structural view of the progressive fluorination approach. The expression system was also used to study the role of Trp68 in the binding of riboflavin by the membrane-bound riboflavin transport protein RibU from . Progressive fluorination of Trp68 revealed a strong electrostatic component that contributed 15-20% to the total riboflavin-RibU binding energy.
Topics: Amino Acids; Cations; Lactococcus lactis; Membrane Transport Proteins; Riboflavin; Tryptophan
PubMed: 35868012
DOI: 10.1021/jacs.2c04986