“…The α-helix content was not altered almost in the presence of SiO 2 NPs, which depicts that the interaction between CAT and SiO 2 NPs does not result in any significant change in the secondary structure of CAT. As deduced from Figure 5, the minimum positions and the ellipticity values of the CD spectra are not changed even after enhancing the molar ratio of NP, evincing that the native structure of CAT remains stable after the interaction.35Figure 5CD spectra of CAT in the presence of different concentrations of SiO 2 NPs: 0 (black), 5 (red), 10 (blue) and 20 μM (green), at room temperature.…”
Section: Resultsmentioning
confidence: 82%
“…The entropy change ( T ∆ S ) was estimated from the intercept to be 81.62±9.11, 84.90±10.28 and 86.27±13.35 kJ/mol at 298, 310 and 315 K, respectively. The positive signs of ∆ H and ∆ S demonstrate that hydrophobic forces are the main contributing interactions in the formation of the SiO 2 NP/CAT complex.35…”
Section: Resultsmentioning
confidence: 93%
“…CD is a potential technique usually employed for the detection of changes in secondary and tertiary conformation of protein 35,36. Backbone configuration of the protein is explored by the far-UV CD ellipticity changes within the range of 190–260 nm.…”
Section: Resultsmentioning
confidence: 99%
“…Backbone configuration of the protein is explored by the far-UV CD ellipticity changes within the range of 190–260 nm. In this range of wavelength, the secondary structure of the protein represents a characteristic magnitudewhich can be denatured or destabilized by triggering the structural alterations during the binding of NPs to the protein.35 In order to explore the change in the secondary structure of CAT in the presence of varying concentrations of SiO 2 NPs (5, 10 and 20 µm), far-UV CD spectra were measured (Figure 5). The CD spectrum of CAT demonstrates two characteristic negative peaks at 208 and 222 nm, indicating the alpha-helical configuration of CAT.…”
Aim
Nanoparticles (NPs) have been receiving potential interests in protein delivery and cell therapy. As a matter of fact, NPs may be used as great candidates in promoting cell therapy by catalase (CAT) delivery into high oxidative stress tissues. However, for using NPs like SiO
2
as carriers, the interaction of NPs with proteins and mesenchymal stem cells (MSCs) should be explored in advance.
Methods
In the present study, the interaction of SiO
2
NPs with CAT and human MSCs (hMSCs) was explored by various spectroscopic methods (fluorescence, circular dichroism (CD), UV-visible), molecular docking and dynamics studies, and cellular (MTT, cellular morphology, cellular uptake, lactate dehydrogenase, ROS, caspase-3, flow cytometry) assays.
Results
Fluorescence study displayed that both dynamic and static quenching mechanisms and hydrophobic interactions are involved in the spontaneous interaction of SiO
2
NPs with CAT. CD spectra indicated that native structure of CAT remains stable after interaction with SiO
2
NPs. UV-visible study also revealed that the kinetic parameters of CAT such as
Km, Vmax, Kcat
, and enzyme efficiency were not changed after the addition of SiO
2
NPs. Molecular docking and dynamics studies showed that Si and SiO
2
clusters interact with hydrophobic residues of CAT and SiO
2
cluster causes minor changes in the CAT structure at a total simulation time of 200 ps. Cellular assays depicted that SiO
2
NPs induce significant cell mortality, change in cellular morphology, cellular internalization, ROS elevation, and apoptosis in hMSCs at higher concentration than 100 µg/mL (170 µM).
Conclusion
The current results suggest that low concentrations of SiO
2
NPs induce no substantial change or mortality against CAT and hMSCs, and potentially useful carriers in CAT delivery to hMSC.
“…The α-helix content was not altered almost in the presence of SiO 2 NPs, which depicts that the interaction between CAT and SiO 2 NPs does not result in any significant change in the secondary structure of CAT. As deduced from Figure 5, the minimum positions and the ellipticity values of the CD spectra are not changed even after enhancing the molar ratio of NP, evincing that the native structure of CAT remains stable after the interaction.35Figure 5CD spectra of CAT in the presence of different concentrations of SiO 2 NPs: 0 (black), 5 (red), 10 (blue) and 20 μM (green), at room temperature.…”
Section: Resultsmentioning
confidence: 82%
“…The entropy change ( T ∆ S ) was estimated from the intercept to be 81.62±9.11, 84.90±10.28 and 86.27±13.35 kJ/mol at 298, 310 and 315 K, respectively. The positive signs of ∆ H and ∆ S demonstrate that hydrophobic forces are the main contributing interactions in the formation of the SiO 2 NP/CAT complex.35…”
Section: Resultsmentioning
confidence: 93%
“…CD is a potential technique usually employed for the detection of changes in secondary and tertiary conformation of protein 35,36. Backbone configuration of the protein is explored by the far-UV CD ellipticity changes within the range of 190–260 nm.…”
Section: Resultsmentioning
confidence: 99%
“…Backbone configuration of the protein is explored by the far-UV CD ellipticity changes within the range of 190–260 nm. In this range of wavelength, the secondary structure of the protein represents a characteristic magnitudewhich can be denatured or destabilized by triggering the structural alterations during the binding of NPs to the protein.35 In order to explore the change in the secondary structure of CAT in the presence of varying concentrations of SiO 2 NPs (5, 10 and 20 µm), far-UV CD spectra were measured (Figure 5). The CD spectrum of CAT demonstrates two characteristic negative peaks at 208 and 222 nm, indicating the alpha-helical configuration of CAT.…”
Aim
Nanoparticles (NPs) have been receiving potential interests in protein delivery and cell therapy. As a matter of fact, NPs may be used as great candidates in promoting cell therapy by catalase (CAT) delivery into high oxidative stress tissues. However, for using NPs like SiO
2
as carriers, the interaction of NPs with proteins and mesenchymal stem cells (MSCs) should be explored in advance.
Methods
In the present study, the interaction of SiO
2
NPs with CAT and human MSCs (hMSCs) was explored by various spectroscopic methods (fluorescence, circular dichroism (CD), UV-visible), molecular docking and dynamics studies, and cellular (MTT, cellular morphology, cellular uptake, lactate dehydrogenase, ROS, caspase-3, flow cytometry) assays.
Results
Fluorescence study displayed that both dynamic and static quenching mechanisms and hydrophobic interactions are involved in the spontaneous interaction of SiO
2
NPs with CAT. CD spectra indicated that native structure of CAT remains stable after interaction with SiO
2
NPs. UV-visible study also revealed that the kinetic parameters of CAT such as
Km, Vmax, Kcat
, and enzyme efficiency were not changed after the addition of SiO
2
NPs. Molecular docking and dynamics studies showed that Si and SiO
2
clusters interact with hydrophobic residues of CAT and SiO
2
cluster causes minor changes in the CAT structure at a total simulation time of 200 ps. Cellular assays depicted that SiO
2
NPs induce significant cell mortality, change in cellular morphology, cellular internalization, ROS elevation, and apoptosis in hMSCs at higher concentration than 100 µg/mL (170 µM).
Conclusion
The current results suggest that low concentrations of SiO
2
NPs induce no substantial change or mortality against CAT and hMSCs, and potentially useful carriers in CAT delivery to hMSC.
“…In order to verify the deduction, the binding constants of HSA-PTAI and HSA-TGB were determined (SI Figure S4). From the modified Stern–Volmer equation: …”
Tiagabine
hydrochloride (TGB) is a clinically frequently used drug
for anticonvulsion and reducing epileptic frequency. Over administration
of TGB could bring about adverse effects, such as speech disorder,
depression, and even suicidal tendencies. Therefore, accessible and
sensitive assay for analysis of TGB becomes an urgent need toward
guiding clinical medication. Here, we present the first report on
fluorescence turn-on detection of TGB in urine testing. In this protocol,
a fluorescent dye, perylene tetracarboxylic acid imide derivative
(PTAI), is found specifically occupying the Sudlow site II of human
serum albumin (HSA) and displays a new phenomenon of binding-induced
quenching (BIQ). In presence of TGB, competitive binding of the TGB
to the site II of HSA will trigger release of PTAI, thus successfully
lighting up the fluorescence of PTAI. This label-free assay enjoys
a broader working range (1–350 μM) and lower detection
limit (0.218 μM) than the traditional liquid chromatography
method and is uninterfered by the miscellaneous in the artificial
urine. The BIQ probe highlights the merits of HSA as a quencher and
a molecular recognition unit, and it opens up a way for studying drug-HSA
interaction mechanism and noninvasive pharmaceutical testing.
Even today, talking about sexual dysfunction largely remains a taboo. Therefore less studies have been recorded and fewer remedies given. Erectile dysfunction (ED) is one of the most commonly treated psychological disorders that leads to major distress, interpersonal limitation and reduces the quality of life and marriage. This study aimed to assess a plant‐derived molecule, yohimbine (Yoh; a β‐carboline indole alkaloid often used for ED treatment) and its potential binding phenomenon with haemoglobin (Hb). Successful binding of Yoh with Hb is evident from spectroscopic and molecular‐docking results. Yoh quenched the fluorescence of Hb efficiently through a static mode. The binding affinity was in the order of 105 M−1 with 1:1 stoichiometry. Thermodynamic analyses concluded that the protein–ligand association was spontaneous and was attributed to entropy‐driven exothermic binding. Nonpolyelectrolytic factor was the core, dominating factor. Structural aspects were deciphered through infrared spectroscopy and computational methods. The giant 3D‐protein moiety was significantly perturbed through drug binding. Hydrophobic forces and hydrogen bonding participation were stipulated by molecular modelling data. This study reveals the detailed interaction pattern and molecular mechanism of Hb–Yoh binding, correlating the structure–function relationship for the first time, and therefore holds enormous importance from the standpoint of rational and efficient drug design and development.
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