Phenotypic Characterization of the Binding of Tetracycline to Human Serum Albumin

Because of the widely usage of the veterinary drug tetracycline (TC), its residue exist extensively in the environment (e.g., animal food, soils, surface water, and groundwater) and can enter human body, being potential harmful. Human serum albumin (HSA) is a major transporter for endogenous and exogenous compounds in vivo. The aim of this study was to examine the interaction of HSA with TC through spectroscopic and molecular modeling methods. The inner filter effect was eliminated to get accurate binding parameters. The site marker competition experiments revealed that TC binds to site II (subdomain IIIA) of HSA mainly through electrostatic interaction, illustrated by the calculated negative ΔH° and ΔS°. Furthermore, molecular docking was applied to define the specific binding sites, the results of which show that TC mainly interacts with the positively charged amino acid residues Arg 410 and Lys 414 predominately through electrostatic force, in accordance with the conclusion of thermodynamic analysis. The binding of TC can cause conformational and some microenvironmental changes of HSA, revealed by UV−visible absorption, synchronous fluorescence, and circular dichroism (CD) results. The accurate and full basic data in the work is beneficial to clarifying the binding mechanism of TC with HSA in vivo and understanding its effect on protein function during the blood transportation process

Noncovalent Interaction of Oxytetracycline with the Enzyme Trypsin

Oxytetracycline (OTC) is a kind of widely used veterinary drugs. The residue of OTC in the environment (e.g., animal food, soils, surface, and groundwater) is potentially harmful. In this article, the binding mode of OTC with trypsin was investigated using spectroscopic and molecular docking methods. OTC can interact with trypsin with one binding site to form OTC−trypsin complex, resulting in inhibition of trypsin activity and change of the secondary structure and the microenvironment of the tryptophan residues of trypsin. After elimination of the inner filter effect, the association constant, K, was calculated to be K290K = 1.36 × 105 L mol−1, K298K = 7.30 × 104 L mol−1, and K307K = 3.58 × 104 L mol−1 at three different temperatures. The calculated thermodynamic parameters (negative values of ΔH○ and ΔS○) indicated that van der Waals interactions and hydrogen bonds play a major role during the interaction. The molecular docking study revealed that OTC bound into the S1 binding pocket, which illustrates that the trypsin activity was competitively inhibited by OTC, in accordance with the conclusion of the trypsin activity experiment. This work establishes a new strategy to probe the toxicity of OTC and contributes to clarify its molecular mechanism of toxicity in vivo. The combination of spectroscopic and molecular docking methods in this work can be applied to investigate the potential enzyme toxicity of other small organic pollutants and drugs

New Insights into the Behavior of Bovine Serum Albumin Adsorbed onto Carbon Nanotubes: Comprehensive Spectroscopic Studies

Bovine serum albumin (BSA) nonspecifically binds to well-dispersed multiwalled carbon nanotubes (MWCNTs), forming a stable bioconjugate. After accounting for the inner filter effect, we found the fluorescence intensity of BSA was quenched by MWCNTs in static mode, which was authenticated by lifetime measurements and Stern−Volmer calculations. The thermodynamic parameters ΔG°, ΔS°, and ΔH° were −9.67 × 103 + 2.48 × 103 ln λ J·mol−1, 41.0 − 0.828 ln λ J·mol−1·K−1, and 7.30 × 103 + 2.23 × 103 ln λ J·mol−1 (λ −4), respectively, which shows a spontaneous and electrostatic interaction. Scatchard analysis and UV−visible results provide statistical data concerning changes in the microenvironment of amide moieties in response to different doses of MWCNTs, revealing different behavior of the BSA molecules. The absorption spectra also show that the tertiary structure of the protein was partially destroyed. The content of secondary structure elements of BSA was changed by the tubes. This work elucidates the interaction mechanism of BSA and MWCNTs from a spectroscopic angle

Binding of Oxytetracycline to Bovine Serum Albumin: Spectroscopic and Molecular Modeling Investigations

The residue of the widely used veterinary drug oxytetracycline (OTC) in the environment (e.g., animal food, soils, surface water, and groundwater) is potentially harmful. Knowledge of its binding to proteins contributes to the understanding of its toxicity in vivo. This work establishes the binding mode of OTC with bovine serum albumin (BSA) under physiological conditions by spectroscopic methods and molecular modeling techniques. The inner filter effect was eliminated to get accurate data (binding parameters). On the basis of the thermodynamic results and site marker competition experiments, it was considered that OTC binds to site II (subdomain IIIA) of BSA mainly by electrostatic interaction. Furthermore, using the BSA model established with CPHmodels, molecular docking and some other molecular modeling methods were applied to further define that OTC interacts with the Arg 433, Arg 436, Ala 429, and Pro 516 residues of BSA. In addition, UV−visible absorption, synchronous fluorescence, and circular dichroism (CD) results showed that the binding of OTC can cause conformational and some microenvironmental changes of BSA. The work provides accurate and full basic data for clarifying the binding mechanisms of OTC with BSA in vivo and is helpful for understanding its effect on protein function during its transportation and distribution in blood

Molecular structure of OTC.

<p>Molecular structure of OTC.</p

MDA content of hRBCs under different OTC concentrations.

<p>Data represent the mean ± SD of three independent experiments.</p

Hemolysis rate (a) and photographs of hemolysis (b) of hRBCs incubated with OTC.

<p>(a) hRBCs incubated with OTC at different concentrations ranging from 1 to 4×10⁻⁴ mol L⁻¹ for 3 h. Data represent the mean ±SD from at least three independent experiments. (b) The presence of red hemoglobin in the supernatant indicates damaged hRBCs. D.I. water (+) and PBS (−) are used as positive and negative control, respectively.</p

Effect of OTC on ATPase activity of hRBCs.

<p>Data represent the mean ± SD of three independent experiments.</p

Effect of OTC on SOD activity of hRBCs.

<p>Data represent the mean ± SD of three independent experiments.</p

Effect of OTC on GSH-Px activity of hRBCs.

<p>Data represent the mean ± SD of three independent experiments.</p