Human serum albumin (HSA) is the most abundant plasma protein in circulation. The three most important drug-binding sites on HSA are Sudlow's Site I (subdomain IIA), Sudlow's Site II (subdomain IIIA), and Heme site (subdomain IB). Heme site and Site I are allosterically coupled; therefore, their ligands may be able to allosterically modulate the binding affinity of each other. In this study, the effects of four Heme site ligands (bilirubin, biliverdin, hemin, and methyl orange) on the interaction of the Site I ligand warfarin with HSA were tested, employing fluorescence spectroscopic, ultrafiltration, and ultracentrifugation studies. Our major results/conclusions are the following. (1) Quenching studies indicated no relevant interaction, while the other fluorescent model used suggested that each Heme site ligand strongly decreases the albumin binding of warfarin. (2) Ultrafiltration and ultracentrifugation studies demonstrated the complex modulation of warfarin-HSA interaction by the different Heme site markers; for example, bilirubin strongly decreased while methyl orange considerably increased the bound fraction of warfarin. (3) Fluorescence spectroscopic studies showed misleading results in these diligand-albumin interactions. (4) Different Heme site ligands can increase or decrease the albumin binding of warfarin and the outcome can even be concentration dependent (e.g., biliverdin and hemin).
- MeSH
- Bilirubin MeSH
- Biliverdine * MeSH
- Heme metabolism MeSH
- Hemin MeSH
- Humans MeSH
- Ligands MeSH
- Serum Albumin metabolism MeSH
- Warfarin * pharmacology MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
The structures of DNA G-quadruplexes are essential for their functions in vivo and in vitro. Our present study revealed that sequential order of the three G-quadruplex loops, that is, loop transposition, could be a critical factor to determinate the G-quadruplex conformation and consequently improved the catalytic function of G-quadruplex based DNAzyme. In the presence of 100mM K+, loop transposition induced one of the G-quadruplex isomers which shared identical loops but differed in the sequential order of loops into a hybrid topology while the others into predominately parallel topologies. 1D NMR spectroscopy and mutation analysis suggested that the hydrogen bonding from loops residues with nucleotides in flanking sequences may be responsible for the stabilization of the different conformations. A well-known DNAzyme consisting of G-quadruplex and hemin (Ferriprotoporphyrin IX chloride) was chosen to test the catalytic function. We found that the loop transposition could enhance the reaction rate obviously by increasing the hemin binding affinity to G-quadruplex. These findings disclose the relations between the loop transposition, G-quadruplex conformation and catalytic function of DNAzyme.
The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE „uniform requirements“ for biomedical papers.
- MeSH
- Acute Disease MeSH
- Hemin therapeutic use MeSH
- Clinical Laboratory Techniques MeSH
- Humans MeSH
- Porphyrias * diagnosis drug therapy therapy MeSH
- Prognosis MeSH
- Check Tag
- Humans MeSH
- Publication type
- Practice Guideline MeSH
- Keywords
- neuroviscerální ataky,
- MeSH
- Adult MeSH
- Heme metabolism MeSH
- Hemin administration & dosage MeSH
- Skin Manifestations MeSH
- Aminolevulinic Acid metabolism MeSH
- Contraindications, Drug MeSH
- Humans MeSH
- Metalloporphyrins metabolism MeSH
- Drug-Related Side Effects and Adverse Reactions MeSH
- Porphyria, Variegate * genetics metabolism physiopathology MeSH
- Photosensitivity Disorders genetics MeSH
- Check Tag
- Adult MeSH
- Humans MeSH
- Male MeSH
- Publication type
- Case Reports MeSH
The genus Bartonella comprises numerous species with at least 13 species pathogenic for humans. They are fastidious, aerobic, Gram negative, and facultative intracellular bacteria which cause a variety of human and non-human diseases. This study focused on the development of a serum-free liquid medium for culture of Bartonella species. Some liquid media are available commercially but all of them use undefined supplements such as fetal calf serum or defibrinated sheep blood. Our intention was to create a reproducible liquid medium for Bartonella species that can simply be prepared. We tested several supplements that could potentially support the growth of Bartonella species. Slight growth improvement was achieved with glucose and sucrose. However, hemin in particular improved the growth rate. At a temperature of 37 °C, a CO2 concentration of 5 %, a humidified atmosphere, and the use of the supplements glucose, sucrose, and hemin, we developed a medium that does not need serum as an undefined supplement any more. In conclusion, the newly developed medium supports growth of Bartonella species equal to the commercially available media but with the advantage that it has a serum-free formulation. It can be prepared fast and easy and is a useful tool in studying these bacteria.
- MeSH
- Bacteriological Techniques methods MeSH
- Bartonella growth & development MeSH
- Glucose metabolism MeSH
- Hemin metabolism MeSH
- Culture Media, Serum-Free chemistry MeSH
- Humans MeSH
- Carbon Dioxide metabolism MeSH
- Sucrose metabolism MeSH
- Temperature MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
Fenomén trpasličích kmenů S. aureus, tzv. Small Colony Variants (SČV), je spojen s chronickými a rekurentními stafylokokovými infekcemi. Tyto fenotypové varianty se liší od kmenů S. aureus s běžným fenotypem nejčastěji velikostí kolonií, jejich morfologií, pigmentací a dalšími znaky, ale také molekulárně genetickými změnami. Příčinou vzniku SCV fenotypu je často mutace v některém z důležitých metabolických či regulačních genů, která je spojena s auxotrofií. Z klinického hlediska je významná zvýšená schopnost SCV kmenů odolávat antibiotické léčbě, zapříčiněná jednak rezistencí k určitým antibiotikům spojenou s příčinou SCV fenotypu a také se schopností těchto kmenů perzistovat uvnitř hostitelských buněk.
The phenomenon of dwarf colonies of S. aureus, the so-called small colony variants (SCVs), is associated with chronic and recurrent staphylococcal infections. Most frequently, these phenotypic variants differ from normal strains of S. aureus in colony size, morphology, pigmentation and other characteristics as well as molecular genetic changes. SCVs frequently emerge as a result of mutations in metabolically important and regulatory genes. The mutations are a cause of SCVs auxotrophy. From a clinical point of view, an increased ability of SCVs to resist antibiotic therapy and also an ability to persist within eukaryotic host cells are of importance.
- MeSH
- Anti-Bacterial Agents therapeutic use MeSH
- Cystic Fibrosis drug therapy microbiology MeSH
- Phenotype MeSH
- Hemin MeSH
- Culture Techniques methods statistics & numerical data trends MeSH
- Humans MeSH
- Molecular Biology MeSH
- Mutation genetics MeSH
- Staphylococcal Infections * diagnosis etiology microbiology MeSH
- Staphylococcus aureus * isolation & purification pathogenicity MeSH
- Vitamin K 3 metabolism MeSH
- Check Tag
- Humans MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
- Review MeSH
The locations of three bilirubin (BR)-binding sites with different affinities were identified as subdomains IB, IIA and IIIA for five mammalian serum albumins (SAs): human (HSA), bovine (BSA), rat, (RSA), rabbit (RbSA) and sheep (SSA). The stereoselectivity of a high-affinity BR-binding site was identified in the BR/SA=1/1 system by circular dichroism (CD) spectroscopy, the sites with low affinity to BR were analyzed using difference CD. Site-specific ligand-competition experiments with ibuprofen (marker for subdomain IIIA) and hemin (marker for subdomain IB) did not reveal any changes for the BR/SA=1/1 system and showed a decrease of the bound BR at BR/SA=3/1. Both sites were identified as sites with low affinity to BR. The correlation between stereoselectivity and the arrangement of Arg-Lys residues indicated similarity between the BR-binding sites in subdomain IIIA for all of the SAs studied. Subdomain IB in HSA, BSA, SSA and RbSA has P-stereoselectivity while in RSA it has M-selectivity toward BR. A ligand-competition experiment with gossypol shows a decrease of the CD signal of bound BR for the BR/SA=1/1 system as well as for BR/SA=3/1. Subdomain IIA was assigned as a high-affinity BR-binding site. The P-stereoselectivity of this site in HSA (and RSA, RbSA) was caused by the right-hand localization of charged residues R257/R218-R222, whereas the left-hand orientation of R257/R218-R199 led to the M-stereoselectivity of the primary binding site in BSA (and SSA).
- MeSH
- Amino Acid Motifs MeSH
- Bilirubin chemistry metabolism MeSH
- Circular Dichroism MeSH
- Hemin chemistry metabolism MeSH
- Ibuprofen chemistry metabolism MeSH
- Rabbits MeSH
- Rats MeSH
- Humans MeSH
- Ligands * MeSH
- Molecular Conformation MeSH
- Sheep MeSH
- Serum Albumin chemistry metabolism MeSH
- Molecular Docking Simulation MeSH
- Cattle MeSH
- Protein Structure, Tertiary MeSH
- Protein Binding MeSH
- Binding Sites MeSH
- Animals MeSH
- Check Tag
- Rabbits MeSH
- Rats MeSH
- Humans MeSH
- Cattle MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Treatment of Helicobacter pylori cells with several chaotropic agents resulted in different degrees of inhibition in the binding of the bacteria to hemin and Congo-red dye. Polyanions also yielded a >50% inhibitory effect. Furthermore, hydrophobic interaction chromatography was used to determine the relative surface hydrophobicity of cell-associated proteins extracted with 3 mol/L urea, revealing proteins with a significant hydrophobic profile.
- MeSH
- Bacterial Adhesion MeSH
- Lithium Chloride pharmacology MeSH
- Sodium Chloride pharmacology MeSH
- Electrophoresis, Polyacrylamide Gel MeSH
- Ethylene Glycol pharmacology MeSH
- Helicobacter pylori drug effects metabolism MeSH
- Hemin metabolism MeSH
- Hydrophobic and Hydrophilic Interactions MeSH
- Hydrogen-Ion Concentration MeSH
- Congo Red metabolism MeSH
- Membrane Proteins metabolism MeSH
- Urea pharmacology MeSH
- Temperature MeSH
- Protein Binding MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- MeSH
- Animal Experimentation MeSH
- Financing, Organized MeSH
- Gangliosides diagnostic use MeSH
- Hemin MeSH
- Cholestasis, Intrahepatic enzymology MeSH
- Disease Models, Animal MeSH
- Rats, Wistar MeSH
- Animals MeSH
- Check Tag
- Female MeSH
- Animals MeSH
- Publication type
- Abstracts MeSH
