A graphene binding-promoted fluorescence enhancement for bovine serum albumin recognition

Chemically converted graphene (CCG) was found to greatly enhance the fluorescence response of squaraine (SQ) dyes to bovine serum albumin (BSA). Addition of BSA to the SQ-CCG solution, where squaraine dyes absorbed on the CCG surface, raised the fluorescence intensity by as much as 80 fold.     

AIE probes towards biomolecules: the improved selectivity with the aid of graphene oxide

Fluorescence probes with aggregation-induced emission(AIE) property have attracted much research interest and have been identified as novel fluorescence biosensors with "turn-on" response. In this mini review, we present an overview of the recent work on improving the selectivity of AIE biosensors with the aid of graphene oxide(GO), according to the displacement strategy.     

Fabrication of a bilayer potentiometric phosphate biosensor by cross-link immobilization with bovine serum albumin and glutaraldehyde

Chemical cross-linking of purine nucleoside phosphorylase (PNP) and xanthine oxidase (XOD) with glutaraldehyde (GLA) and bovine serum albumin (BSA) has been used to fabricate a stable and reliable bilayer potentiometric phosphate biosensor. The bilayer arrangement consists of an inner BSA-GLA layer and an outer BSA-GLA-PNP-XOD layer. The inclusion of the inner BSA-GLA layer improves the adhesion of the outer BSA-GLA-PNP-XOD layer and ensures stability of the phosphate biosensor. Established optimum conditions for immobilization of the enzymes in the outer layer and for reliable potentiometric measurement were 4.5% v/v GLA, 6.8% w/v BSA, XOD:PNP mole ratio of 1:8, and a film drying time of 30 min. As little as 20 μM of phosphate can be detected with the BSA-GLA/BSA-GLA-XOD-PNP bilayer biosensor with a linear concentration range between 40 and 120 μM. The biosensor was very stable for 21 days, achieving a good reproducibility with a rsd of only 5.7% and, even after more than a month, the change in the initial potential value was only 10%.     

Binding of naproxen to bovine serum albumin and tryptophan-modified bovine serum albumin

Two classes of binding sites, a single high-affinity site with an association constant of 4·8×10⁶ M⁻¹ and two low-affinity sites with association constant of about 0·05×10⁶ M⁻¹ have been observed in the interaction of Naproxen with bovine serum albumin (BSA). Chemical modification of two tryptophan residues in BSA with 2-hydroxy-5-nitrobenzyl bromide has led to a reduction in the association constant of the high-affinity site by 89% and its number of binding sites by 66% suggesting the involvement of tryptophan residues in the high-affinity site. In contrast, the two low-affinity sites were not affected by the modification. Binding of Naproxen to the low-affinity sites of BSA induces microdisorganisation of the albumin structure leading to conformational changes as evident from fluorescence measurements with 1-anilino-8-naphthalenesulphonic acid as the probe.     

Porous graphene oxide-based carbon artefact with high capacity for methylene blue adsorption

A three-dimensional porous graphene oxide (PGO) material prepared by hydrothermal method was selected to adsorb methylene blue (MB), which demonstrates a high MB adsorption capacity, up to 1100 mg g^?1 in alkaline solution at room temperature. The influences of different pore structures and different contents of oxygen-containing functional groups on MB adsorption behaviors were studied in detail, which indicated that the high MB adsorption capacity is mainly ascribed to the synergistic effect of the large number of oxygen-containing functional groups and the interconnected 3D porous network. Moreover, based on the investigation on the adsorption kinetics and the effect of pH value on MB adsorption, we propose a two-step adsorption kinetics for PGO, which involved in two interactions between MB molecular and porous graphene oxide-based carbon: electrostatic force and π-π stacking. Besides, the calculation of the activation energies indicates that chemisorption dominates the adsorption for PGO in comparison with physisorption for three-dimensional porous graphene materials which has low adsorption capacity because of the removal of functional groups. The results are of great significance for the design and environmental applications of PGO as a promising adsorbent material for water purification.     

硫堇-牛血清白蛋白氧化还原复合膜修饰葡萄糖生物传感器的研究

通过交联法和自组装法制备了一种双酶型葡萄糖生物传感器.首先以牛血清白蛋白-戊二醛为交联剂以实现对辣根过氧化物酶(HRP)的固载,再利用凝集素-糖蛋白的识别作用将葡萄糖氧化酶(GOD)分子组装到电极表面,制得双酶型的葡萄糖生物传感器.采用原子力显微镜(AFM)考察了复合膜的性质,同时采用循环伏安法和计时电流法考察了该传感...     

A facile graphene oxide-based DNA polymerase assay

The DNA polymerase assay is fundamental for related molecular biology investigations and drug screenings, however, the commonly used radioactive method is laborious and restricted. Herein, we report a novel, simple and cost-effective fluorometric DNA polymerase detection method by utilizing graphene oxide (GO) as a signal switch. In this strategy, in the absence of DNA polymerase, the fluorophore-labeled template ssDNA could be strongly adsorbed and almost entirely quenched by GO. However, as DNA polymerase exists, the polymerized dsDNA product might lead to a much lower quenching efficiency after addition of GO due to the much weaker interaction of dsDNA with GO than ssDNA, thus resulting in a much higher fluorescence signal detected. As proof of concept, the quantitative DNA polymerase activity assay was performed using the Klenow fragment exo(-) (KF(-)) as a model. It was confirmed that, after optimization of detection conditions, KF(-) activity could be sensitively detected through facile fluorescence measurements, with a detection limit of 0.05 U mL(-1) and a good linear correlation between 0.05-2.5 U mL(-1) (R(2) = 0.9928). In addition, this GO-based method was further inspected to evaluate the inhibitive behaviors of several drugs toward KF(-) activity, the result of which firmly demonstrated its potential application in polymerization-targeted drug screening.     

间苯二酚与牛血清白蛋白的作用机理

采用荧光和紫外光谱法研究了间苯二酚与牛血清白蛋白(BSA)的相互作用。间苯二酚使BSA的构象发生改变,α-螺旋含量减小。同步荧光光谱发现间苯二酚使BSA色氨酸残基的疏水性降低,酪氨酸残基的疏水性增强。荧光光谱表明猝灭机理为静态猝灭,计算了复合物的结合常数,通过热力学参数得出间苯二酚与BSA之间的作用力主要是静电作用力。     

邻苯二酚与牛血清白蛋白的相互作用

采用荧光和紫外光谱法研究了邻苯二酚同牛血清白蛋白(BSA)的作用.邻苯二酚对BSA内源荧光猝灭机理为静态猝灭,两者之间生成了不发荧光的复合物.根据双对数方程计算了复合物的结合常数KA和结合位点数n.确定邻苯二酚与BSA只有一个结合位点(可能位于Site I).通过热力学参数得出邻苯二酚与BSA之间主要作用力是疏水作用.同步荧光的结果表明邻苯二酚改变了BSA的分子构象,使色氨酸残基的极性增加,酪氨酸残基的疏水性增强.     

A graphene oxide-based nano-beacon for DNA phosphorylation analysis

We designed a single-fluorophore-tagged hairpin-structured nano-beacon probe by using a superquencher, graphene oxide (GO), based on which a new method for the analysis of DNA phosphorylation detection was developed.     

A lateral flow biosensor for detection of nucleic acids with high sensitivity and selectivity

A lateral flow biosensor based on isothermal strand-displacement polymerase reaction and gold nanoparticles has been developed for the visual detection of nucleic acids with a detection limit of 0.01 fM.     

Binding interaction of gatifloxacin with bovine serum albumin.

The binding of gatifloxacin to bovine serum albumin (BSA) in aqueous solution was studied using fluorescence spectroscopy and absorbance spectra, Further, the interactions influenced by Fe3+ and Cu2+ were also explored in this work. Based on Scatchard's site-binding model and florescence quenching, practical formulas for small molecule ligands to bio-macromolecules have been proposed. The binding parameters were measured according to suggested models, and the binding distance and the transfer efficiency of energy between gatifloxacin and BSA were also obtained in view of the F?rster theory of non-radiation energy transfer. The effect of gatifloxacin on the conformation of BSA has also been analyzed using synchronous fluorescence spectroscopy.     

Binding of chloroquine-conjugated gold nanoparticles with bovine serum albumin

We have conjugated chloroquine, an anti-malarial, antiviral and anti-tumor drug, with thiol-functionalized gold nanoparticles and studied their binding interaction with bovine serum albumin (BSA) protein. Gold nanoparticles have been synthesized using sodium borohydride as reducing agent and 11-mercaptoundecanoic acid as thiol functionalizing ligand in aqueous medium. The formation of gold nanoparticles was confirmed from the characteristic surface plasmon absorption band at 522 nm and transmission electron microscopy revealed the average particle size to be ~7 nm. Chloroquine was conjugated to thiolated gold nanoparticles by using EDC/NHS chemistry and the binding was analyzed using optical density measurement and Fourier transform infrared spectroscopy. The chloroquine-conjugated gold nanoparticles (GNP-Chl) were found to interact efficiently with BSA. Thermodynamic parameters suggest that the binding is driven by both enthalpy and entropy, accompanied with only a minor alteration in protein's structure. Competitive drug binding assay revealed that the GNP-Chl bind at warfarin binding site I in subdomain IIA of BSA and was further supported by Trp212 fluorescence quenching measurements. Unraveling the nature of interactions of GNP-Chl with BSA would pave the way for the design of nanotherapeutic agents with improved functionality, enriching the field of nanomedicine.     

18β-Glycyrrhetinic acid interaction with bovine serum albumin

The interaction of 18β-glycyrrhetinic acid (GA): The metabolite of glycyrrhizic acid which is the main active component of a commonly used traditional Chinese medicine (TCM) Glycyrrhiza Uralensis Fisch with bovine serum albumin (BSA) has been investigated. Fluorescence emission spectra of serum albumin in the presence of GA, recorded at the excitation wavelength 280 nm, clearly show that GA act as quencher and have different quenching mechanism at a pH below or above the isoelectric point (pI). The binding sites number n and apparent binding constant K were measured. The thermodynamic parameters ΔH°, ΔG°, ΔS° at different temperatures were calculated. The effects of some common metal ions on binding are considered. Synchronous fluorescence and UV–vis spectra were used to study protein conformation. Energy transfer between GA and HSA was calculated by Förster's theory and the binding site was suggested to be site II. The binding of monoammonium glycyrrhizinate (GL) to BSA is also compared.     

Interaction of polyethyleneimine-functionalized ZnO nanoparticles with bovine serum albumin

In biological fluids, nanoparticles are always surrounded by proteins. As the protein is adsorbed on the surface, the extent of adsorption and the effect on the protein conformation and stability are dependent on the chemical nature, shape, and size of the nanoparticle (NP). We have carried out a detailed investigation on the interaction of bovine serum albumin (BSA) with polyethyleneimine-functionalized ZnO nanoparticles (ZnO-PEI). ZnO-PEI was synthesized using a wet chemical method with a core size of ~3-7 nm (from transmission electron microscopy). The interaction of BSA with ZnO-PEI was examined using a combination of calorimetric, spectroscopic, and computational techniques. The binding was studied by ITC (isothermal titration calorimetry), and the result revealed that the complexation is enthalpy-driven, indicating the possible involvement of electrostatic interaction. To investigate the nature of the interaction and the location of the binding site, a detailed domain-wise surface electrostatic potential calculation was performed using adaptive Poisson-Boltzmann software (APBS). The result shows that the protein surface can bind the nanoparticle. On binding ZnO-PEI, the protein gets destabilized to some extent, as displayed by CD (circular dichroism) and FTIR (Fourier transform infrared) spectroscopy. Chemical and thermal denaturation of BSA, when carried out in the presence of ZnO-PEI, also indicated a small perturbation in the protein structure. A comparison of the enthalpy and entropy components of binding with those derived for the interaction of BSA with ZnO nanoparticles explains the effect of hydrophilic cationic species attached on the NP surface. The effect of the NP surface modification on the structure and stability of BSA would find useful applications in nanobiotechnology.     

Hydrophobic interactions of phenoxazine modulators with bovine serum albumin

The interaction of 10-(3′-N-morpholinopropyl)phenoxazine [MPP], 10-(4′-N-morpholinobutyl)phenoxazine [MBP], 10-(3′-N-morpholinopropyl)-2-chlorophenoxazine [MPCP], 10-(3′-N-piperidinopropyl)-2-chlorophenoxazine [PPCP] or 10-(3′-N-morpholinopropyl)-2-trifluoromethylphenoxazine [MPTP] with bovine serum albumin (BSA) has been studied by gel filtration and equilibrium dialysis methods. The binding of these modulators, based on dialysis experiments, has been characterized using the following parameters: percentage of bound drug (Β), the association constant (K ₁), the apparent binding constant (k) and the free energy change (δF‡). The binding of phenoxazine derivatives to serum transporter protein, BSA, is correlated with their octanol-water partition coefficient, log₁₀ P. In addition, effect of the displacing activities of hydroxyzine and acetylsalicylic acid on the binding of phenoxazine derivatives to albumin has been studied. Results of the displacement experiments show that phenoxazine benzene rings and tertiary amines attached to the side chain of the phenoxazine moiety are bound to a hydrophobic area on the albumin molecule.     

Pulsed-electric-field crossflow ultrafiltration of bovine serum albumin

A conventional crossflow ultrafiltration (CUF) apparatus was modified by the inclusion of electrodes which permitted a pulsed electric field to be produced across the ultrafiltration membrane (PEF-UF process). Using this apparatus, a discontinuous electrophoretic velocity was imposed upon the proteins being concentrated, opposing their convective movement toward the CUF membrane. This resulted in a lower concentration of rejected solute protein in the fluid boundary layer adjacent to the high-pressure side of the membrane and, hence, in a lower solute-related filtration resistance than in the case of conventional ultrafiltration (zero electric field). Studies of the PEF-UF process with bovine serum albumin (BSA) in the range of 0.5–5% w/v demonstrated a 25–40% decrease in the solute-related resistance to the permeate flux compared to the case of a zero electric field. Accordingly, higher permeate fluxes and, therefore, higher rates of concentration of the protein solution were obtained than for conventional crossflow ultrafiltration. When the electric field was reimposed following a period of operation under conventional CUF conditions, the permeate flux could be restored to nearly the same higher value observed initially for the PEF-UF process.