Study on the multiple sites binding of human serum albumin and porphyrin by affinity capillary electrophoresis

Equations to describe the two sites binding between proteins and ligands were deduced. According to these equations, not only the binding constants, but also the mole fraction of proteins in different forms could be obtained. Using the published data on the interaction between human serum albumin (HSA) and three kinds of porphyrin (coproporphyrin (CP), uroporphyrin I (UP) and protoporphyrin (PP)), a further study on their binding was carried out. It was concluded that there may exist two binding sites with the binding constants at the first site. proved to be the preferential one, being 6.50 x 10(5) 1.94 x 10(6) and 8.94 x 10(5). respectively. In addition. it was also demonstrated that the two binding sites of HSA with CP and UP might be of different kinds, though those of HSA and PP were of the same kind but at different positions. (C) 2002 Elsevier Science B.V. All rights reserved.

Binding of metal ions with protein studied by a combined technique of microdialysis with liquid chromatography

A method has been developed for the determination of interactions of metal ions and protein by using microdialysis sampling technique combined with pre-column derivation and reversed-phase ion-pair liquid chromatographic (HPLC analysis. Cu(II), Zn(II) and human serum albumin (HSA) were chosen as model metal ions and protein, respectively. The mixed solutions of metal ions and HSA with different molar ratios buffered with 0.1 M Tris-HCl containing 0.1 M NaCl at pH 7.43 were sampled with a mirodialysis probe by keeping perfusion rate at 1 mul/min and the temperature at 37 degreesC. The free concentrations of metal ions in microdialysates were assayed by precolumn derivatization with meso-tetra(4-sulfophenyl)-porphyrin (TPPS4) followed ion-pair HPLC analysis. The recovery (R) of microdialysis sampling was measured in vitro under similar conditions as 65.74% for Cu(II), 70.45% for Zn(II) with R.S.D. below 3.2%. The primary binding constants and number of binding site estimated by the Scatchard plot analysis are 5.04 x 10(6) M-1 and 0.85 for Cu(II), and 9.87 x 10(6) M-1 and 1.10 for Zn(II), respectively. The competition of Cu(II) and Zn(II) at the second binding site on HSA was investigated, and it was observed that there is a second site on HSA to bind Cu(II) and Zn(II), the affinity of Cu(II) is stronger than that of Zn(II) to this second site of HSA. (C) 2001 Elsevier Science B.V. All rights reserved.

Competing binding of metal ions with protein studied by microdialysis

A method has been established to study the competing binding of metal ions with protein by a combined technique of microdialysis with high performance liquid chromatography (HPLC). Ni2+, Cd2+, Zn2+, Cu2+ and human serum albumin (HSA) were chosen as model metal ions and protein. The experimental results show that Ni2+ and Cu2+ share a common primary binding site on HSA, and Zn2+ and Cd2+ share a different common primary binding site from them, but there is a common multi-metal binding site for all of those four metal ions. This method show advantages of fast sampling, easily to be operated and especially to be useful when ideal spectroscopic probes are not available for the study of interaction between protein and metal ions.

Synthesis and Metal-ion Binding Properties of New N2S4O2- and N2S5O2-Donor Macrocycles

Synthetic procedures for new mixed-donor macrocycle compounds were reported. The macrocyclic compounds were used in solvent extraction metal picrates such as Ag+, Hg2+, Cd2+, Zn2+, Cu2+, Ni2+, Mn2+, Pb2+, and Co2+. The metal picrate extractions were investigated at 25±0.1°C with the aid of UV-visible spectrometry. It was found that 6,7,9,10,12,13,23,24-octahydro-19H,26Hdibenzo[h,t](1,4,7,13,16,22,10,19) dioxatetrathiadiazasiclotetracosine-20,27(21H,28H)-dione showed selectivity towards Ag+, Hg2+, and Cd2+ among the other metals. The extraction constants (Log Kex) and complex compositions were determined for the Ag+ and Hg2+ complexes for this compound and 9,10,12,13,23,24,26,27,29,30-decahydro-5H,15H-dibenzo-[h,w][1,4,7,13,16,19,25-,10,22] dioxapentathiadiazacycloheptacosine-6,16(7H,17H)-dione.

Evaluation of flavonoids binding to DNA duplexes by electrospray ionization mass spectrometry

In this study, electrospray ionization mass spectrometry (ESI-MS) was used to investigate the binding interactions of ten flavonoid aglycones and ten flavonoid glycosides with DNA duplexes. Relative binding affinities of the flavonoids toward DNA duplexes were estimated based on the fraction of bound DNA. The results revealed that the 4'-OH group of flavonoid aglycones was essential for their DNA-binding properties. Flavonoid glycosides with sugar chain linked on ring A or ring B showed enhanced binding toward the duplexes over their aglycone counterparts, whereas glycosylation of the flavonol quercetin on ring C exhibited a less pronounced effect.

Studies on alkaloids binding to GC-rich human survivin promoter DNA using positive and negative ion electrospray ionization mass spectrometry

Electrospray ionization mass spectrometry (ESI-MS) was used to investigate the binding of 13 alkaloids to two GC-rich DNA duplexes which are critical sequences in human survivin promoter. Negative ion ESI-MS was first applied to screen the binding of the alkaloids to the duplexes. Six alkaloids (including berberine, jatrorrhizine, palmatine, reserpine, berbamine, and tetrandrine) show complexation with the target DNA sequences. Relative binding affinities were estimated from the negative ion ESI data, and the alkaloids show a binding preference to the duplex with higher GC content. Positive ion ESI mass spectra of the complexes were also recorded and compared with those obtained in negative ion mode.

Microarray-Based Study of Carbohydrate-Protein Binding by Gold Nanoparticle Probes

In order to develop a novel high-throughput tool for monitoring carbohydrate-protein interactions, we prepared carbohydrate or glycoprotein microarrays by immobilizing amino modified carbohydrates on aldehyde-derivatized glass slides or glycoprotein on epoxide-derivatized glass slides and carried out lectin binding experiments by using these microarrays, respectively. The interaction events are marked by attachment of gold nanoparticles followed by silver deposition for signal enhancement. The attachment of the gold nanoparticles is achieved by standard avidin-biotin chemistry.

Single-walled carbon nanotubes binding to human telomeric i-motif DNA under molecular-crowding conditions: More water molecules released

The natural occurrence of the human telomeric G-quadruplex or i-motif in vivo has not been demonstrated and the biological effects of the induction of these structures need to be clarified. Intracellular environments are highly crowded with various biomolecules and in vitro studies under molecular-crowding conditions will provide important information on how biomolecules behave in cells. Here we report that cell-mimic crowding can increase i-motif stability at acid pH and cause dehydration.

Multiplex binding modes of toluidine blue with calf thymus DNA and conformational transition of DNA revealed by spectroscopic studies

It is noteworthy to understand the details of interactions between antitumor drugs and DNA because the binding modes and affinities affect their antitumor activities. Here, The interaction of toluidine blue (TB), a potential antitumor drug for photodynamic therapy of tumor, with calf thymus DNA (ctDNA) was explored by UV-vis, fluorescence, circular dichroism (CD) spectroscopy, UV-rnelting method and surface-enhance Raman spectroscopy (SERS). The experimental results suggest that TB could bind to ctDNA via both electrostatic interaction and partial intercalation.

7-Amino actinomycin D bound to single-stranded hairpin DNA enhanced by loop sequence-dependent luminescent Eu3+ and Tb3+ binding

Lanthanide Eu3+ and Tb3+ ions have been widely used in luminescent resonance energy transfer (LRET) for bioassays to study metal binding microenvironments. We report here that Eu3+ or Tb3+ can increase the binding affinity of antitumor antibiotic drug agent, 7-amino actinomycin D (7AACTD), binding to 5'-GT/TG-5' or 5'-GA/AG-5' mismatched stem region of the single-stranded hairpin DNA. Further studies indicate that the effect of Eu3+ or Tb3+ on 7AACTD binding is related to DNA loop sequence. Our results will provide new insights into how metal ions can enhance antitumor agents binding to their targets.

Single-walled carbon nanotubes binding to human telomeric i-motif DNA: significant acceleration of S1 nuclease cleavage rate

Single-walled carbon nanotubes (SWNTs) binding to human telomeric i-motif DNA can significantly accelerate S1 nuclease cleavage rate by increasing the enzyme turnover number.

Daunomycin binding to detergent micelles: A model system for evaluating the hydrophobic contribution to drug-DNA interactions

The interaction of daunomycin with sodium dodecyl sulfate and Triton X-100 micelles was investigated as a model for the hydrophobic contribution to the free energy of DNA intercalation reactions. Measurements of visible absorbance, fluorescence lifetime, steady-state fluorescence emission intensity, and fluorescence anisotropy indicate that the anthraquinone ring partitions into the hydrophobic micelle interior. Fluorescence quenching experiments using both steady-state and lifetime measurements demonstrate reduced accessibility of daunomycin in sodium dodecyl sulfate micelles to the anionic quencher iodide and to the neutral quencher acrylamide. Quenching of daunomycin fluorescence by iodide in Triton X-100 micelles was similar to that seen with free daunomycin. Studies of the energetics of the interaction of daunomycin with micelles by fluorescence and absorbance titration methods and by isothermal titration calorimetry in the presence of excess micelles revealed that association with sodium dodecyl sulfate and Triton X-100 micelles is driven by a large negative enthalpy. Association of the drug with both types of micelles also has a favorable entropic contribution, which is larger in magnitude for Triton X-100 micelles than for sodium dodecyl sulfate micelles.

Downhill kinetics of biomolecular interface binding: Globally connected scenario

We study the kinetics of the biomolecular binding process at the interface using energy landscape theory. The global kinetic connectivity case is considered for a downhill funneled energy landscape. By solving the kinetic master equation, the kinetic time for binding is obtained and shown to have a U-shape curve-dependence on the temperature. The kinetic minimum of the binding time monotonically decreases when the ratio of the underlying energy gap between native state and average non-native states versus the roughness or the fluctuations of the landscape increases. At intermediate temperatures,fluctuations measured by the higher moments of the binding time lead to non-Poissonian, non-exponential kinetics. At both high and very low temperatures, the kinetics is nearly Poissonian and exponential.