scholarly journals Interaction of polyethylene glycol with cytochrome c investigated via in vitro and in silico approaches

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Zahoor Ahmad Parray ◽  
Faizan Ahmad ◽  
Mohamed F. Alajmi ◽  
Afzal Hussain ◽  
Md. Imtaiyaz Hassan ◽  
...  
Keyword(s):  
Polyethylene Glycol ◽  
Structural Changes ◽  
Apoptotic Cell ◽  
Titration Calorimetry ◽  
Spectroscopic Techniques ◽  
Tertiary Structures ◽  
Time Resolved ◽  
Soft Interactions ◽  
And Control

AbstractOne of the significant proteins that have attracted research groups due to virtue of being a potent selective anticancer drug target and property of triggering apoptosis upon release in cytoplasm is cytochrome c (cyt c). The mechanical transformations due to the macromolecular crowding in membrane in the mammalian cell are proposed to be useful inductors of changes in volume. It is very interesting to know that mitochondrial function were observed to be improved by polyethylene glycol (PEG) interaction, which in turn inhibits the cyt c (a pro-apoptotic cell death factor). In this work, the effect of polyethylene glycol of molecular weight 4 kilo Dalton (PEG 4 kDa) was investigated to highlight the structural transformations (tertiary and secondary structure) in cyt c using a choice of spectroscopic techniques (including UV–Vis absorption, near-UV, far-UV and Soret circular dichroism and fluorescence spectroscopy), which shows noteworthy shifts in the secondary and tertiary structures at higher concentrations of PEG 4 kDa with small changes in the heme-globular interactions. The size distribution changes of native protein treated with various concentrations of the crowder were observed and analyzed by dynamic light scattering (DLS). The interaction studies of the crowder with the protein was observed and analyzed by FTIR, isothermal titration calorimetry, time resolved fluorescence and molecular docking. The investigations suggested that the structural changes in the protein occurred due to soft interactions of PEG 4 kDa, which usually destabilizes proteins. The experimental evidence in this study proposed that crowding could be another approach to mechanical super-competition and free of certain markers that could aid in the identification and control of various diseases. This study suggests that crowders at specific concentrations, which softly interact with proteins, can be exploited as remedy for various diseases.

scholarly journals Synthesis, Characterization and DNA-Binding Affinity of a New Zinc(II) Bis(5-methoxy-indol-3-yl)propane-1,3-dione Complex

2021 ◽  
Vol 14 (8) ◽  
pp. 760
Author(s):  
Luca Scapinello ◽  
Guglielmo Vesco ◽  
Luca Nardo ◽  
Angelo Maspero ◽  
Federico Vavassori ◽  
...  
Keyword(s):  
Metal Ion ◽  
Structural Changes ◽  
Fluorescence Emission ◽  
Spectroscopic Techniques ◽  
Time Resolved ◽  
Fluorescence Studies ◽  
High Affinity ◽  
Drug Substances ◽  
Calf Thymus

The novel zinc(II) µ-oxo-bridged-dimeric complex [Zn2(µ-O)2(BMIP)2] (BMIP = 1,3-bis(5-methoxy-1-methyl-1H-indol-3-yl)propane-1,3-dione), 1, was synthetized and fully characterized. The spectral data indicate a zincoxane molecular structure, with the BMIP ligand coordinating in its neutral form via its oxygen atoms. Structural changes in 1 in dimethylsulfoxide (DMSO) were evidenced by means of spectroscopic techniques including infrared absorption and nuclear magnetic resonance, showing DMSO entrance in the coordination sphere of the metal ion. The resulting complex [Zn2(µ-O)2(BMIP)2(DMSO)], 2, readily reacts in the presence of N-methyl-imidazole (NMI), a liquid-phase nucleoside mimic, to form [Zn2(µ-O)2(BMIP)2(NMI)], 3, through DMSO displacement. The three complexes show high thermal stability, demonstrating that 1 has high affinity for hard nucleophiles. Finally, with the aim of probing the suitability of this system as model scaffold for new potential anticancer metallodrugs, the interactions of 1 with calf thymus DNA were investigated in vitro in pseudo-physiological environment through UV-Vis absorption and fluorescence emission spectroscopy, as well as time-resolved fluorescence studies. The latter analyses revealed that [Zn2(µ-O)2(BMIP)2(DMSO)] binds to DNA with high affinity upon DMSO displacement, opening new perspectives for the development of optimized drug substances.

Influence of Ascorbic Acid on the Structure and Function of Alpha-2- macroglobulin: Investigations using Spectroscopic and Thermodynamic Techniques

2020 ◽  
Vol 27 (3) ◽  
pp. 201-209
Author(s):  
Syed Saqib Ali ◽  
Mohammad Khalid Zia ◽  
Tooba Siddiqui ◽  
Haseeb Ahsan ◽  
Fahim Halim Khan
Keyword(s):  
Ascorbic Acid ◽  
Conformational Changes ◽  
Structural Changes ◽  
The Body ◽  
Titration Calorimetry ◽  
Spectroscopic Techniques ◽  
Human Beings ◽  
Plasma Ascorbic Acid ◽  
Dietary Antioxidant ◽  
And Function

Background: Ascorbic acid is a classic dietary antioxidant which plays an important role in the body of human beings. It is commonly found in various foods as well as taken as dietary supplement. Objective: The plasma ascorbic acid concentration may range from low, as in chronic or acute oxidative stress to high if delivered intravenously during cancer treatment. Sheep alpha-2- macroglobulin (α2M), a human α2M homologue is a large tetrameric glycoprotein of 630 kDa with antiproteinase activity, found in sheep’s blood. Methods: In the present study, the interaction of ascorbic acid with alpha-2-macroglobulin was explored in the presence of visible light by utilizing various spectroscopic techniques and isothermal titration calorimetry (ITC). Results: UV-vis and fluorescence spectroscopy suggests the formation of a complex between ascorbic acid and α2M apparent by increased absorbance and decreased fluorescence. Secondary structural changes in the α2M were investigated by CD and FT-IR spectroscopy. Our findings suggest the induction of subtle conformational changes in α2M induced by ascorbic acid. Thermodynamics signatures of ascorbic acid and α2M interaction indicate that the binding is an enthalpy-driven process. Conclusion: It is possible that ascorbic acid binds and compromises antiproteinase activity of α2M by inducing changes in the secondary structure of the protein.

scholarly journals Conventional frequency ultrasound detection of tumor response in vivo to cancer treatment administration

2021 ◽  
Author(s):  
Naum Papanicolau
Keyword(s):  
Cell Death ◽  
Cancer Therapy ◽  
Structural Changes ◽  
Apoptotic Cell ◽  
High Frequency Ultrasound ◽  
Patient Response ◽  
Tumor Morphology ◽  
Frequency Ultrasound

Current methods employed to evaluate patient response to cancer therapy are typically invasive requiring examination of excised tissue. The development of a non-invasive method of monitoring patient response to cancer therapy administration would potentiate clinical decisions permitting clinicians to adjust therapy regimens early in a treatment course based upon individual patient responses. It has been previously demonstrated that high frequency ultrasound is capable of reliably quantifying structural changes in tumor morphology in response to cancer therapies. Preliminary work has also indicated that ultrasound employed at clinically relevant frequencies (1-15 MHz) can detect apoptotic cell death using in vitro models. This thesis examines changes in tumor morphology in response to cancer therapy administration employing ultrasound at a clinically applicable frequency in a preclinical in vivo mouse model. The power spectrum of the radiofrequency data obtained from tumors was analyzed via linear regression spectroscopic analysis, as well as evaluating a statistical analysis of the amplitude distribution of the signal envelope. It is demonstrated here for the first time that 7 MHz ultrasound can detect apoptotic and other forms of cell death in vivo. A potential for a parametric imaging technique to visually represent analysis results is also demonstrated.

scholarly journals Spectroscopic and In Vitro Investigations of Boron(III) Complex with Meso-4-Methoxycarbonylpropylsubstituted Dipyrromethene for Fluorescence Bioimaging Applications

Molecules ◽  
2020 ◽  
Vol 25 (19) ◽  
pp. 4541
Author(s):  
Galina Guseva ◽  
Elena Antina ◽  
Mikhail Berezin ◽  
Svetlana Lisovskaya ◽  
Roman Pavelyev ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Structural Changes ◽  
Apoptotic Cell ◽  
High Quantum Yield ◽  
Cell Organelles ◽  
Fluorescent Marker ◽  
X Ray ◽  
Fluorescence Bioimaging ◽  
Mycotic Infections

This study focuses on the behavior of a new fluorescent marker for labeling individual biomolecules and staining cell organelles developed on a meso-substituted BODIPY platform. Boron(III) complex with meso-4-methoxycarbonylpropylsubstituted 3,3’,5,5’-tetramethyl-2,2′-dipyrromethene has been synthesized and identified via visible, UV-, NMR- and MS-spectra X-ray. The behavior of fluorophore in solutions has been studied with various experimental techniques. It has been found that luminophore exhibits a high quantum yield (almost ~100–75%) in the blue-green region (513–520 nm) and has high photostability. In addition, biological analysis indicates that the fluorophore exhibits a tendency to effectively penetrate into cell membranes. On the other hand, the proposed BODIPY can be used to study the significant differences among a large number of pathogens of mycotic infections, as well as to visualize structural changes in the plasma membrane, which is necessary for the clearance of mammalian cells undergoing apoptotic cell death.

Filamin A Binding Stabilizes Nascent Glycoprotein Ibα Trafficking and Thereby Enhances Its Surface Expression.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3656-3656
Author(s):  
Shuju Feng ◽  
Xin Lu ◽  
Michael H. Kroll
Keyword(s):  
Cho Cells ◽  
Protein Trafficking ◽  
Structural Changes ◽  
Extracellular Domain ◽  
Surface Expression ◽  
Cytoplasmic Domain ◽  
Filamin A ◽  
Von Willebrand ◽  
And Control

Abstract The glycoprotein (Gp) Ib-IX-V complex is essential for platelet function. The extracellular domain of GpIbα binds to von Willebrand factor (VWF) and rapidly effects shear-dependent platelet rolling, tether formation, adherence and aggregation onto freshly exposed subendothelium. Following VWF binding, GpIbα transduces proaggregatory signals through its cytoplasmic domain, and these signals may be modulated by the activity of GpIbβ and GpV. The expression of GpIbα, GpIbβ and GpIX in megakaryocytes also regulates terminal maturation and platelet release (its absence results in Bernard-Soulier syndrome) and there is human genetic and in vitro evidence that the cytoplasmic domain of GpIbα regulates megakaryocyte growth and surface expression of the complex. The cytoplasmic domain of GpIbα possesses a binding region for filamin A, which links GpIb-IX-V to the platelet cytoskeleton, and there is evidence that filamin A binding to GpIbα directs the surface expression of GpIb-IX in CHO cells. To investigate the mechanism of this effect, we examined GpIbα biosynthesis in CHO cells stably co-expressing wild-type or mutant GpIbα with GpIbβ, GpIX and filamin A. We observed that surface GpIbα expression as measured by flow cytometry is enhanced ~ 1 log-order in CHO cells co-expressing human filamin A. In comparison with CHO-GpIbαβIX cell lysates, lysates from CHO-GpIbαβIX-filamin A cells showed greater amounts of immature (~ 65 kDa), incompletely glycosylated (~ 80 kDa and ~ 90 kDa) and fully mature GpIbα (~ 120 kDa), but lesser amounts of the ~ 15 kDa C-terminal peptide released when the extracellular domain of GpIbα (glycocalycin) is cleaved by surface proteases. To determine if the effect of filamin A is due to its binding to GpIbα, we examined GpIbα biosynthesis using several mutants of GpIbα co-expressed with GpIbβ, GpIX and filamin A. When filamin A binding is eliminated by truncation of GpIbα at C-terminal residue 557 or by a deletion in GpIbα between amino acids 542–570, the decreased synthesis of mature GpIbα is accompanied by nearly complete elimination of all immunodetectable immature GpIbα and by increased immunodetectable C-terminal peptide. To corroborate these data and control for cell selection and non-specific secondary structural changes, we examined the expression of three cell lines expressing two additional mutants of GpIbα in CHO cells stably transfected with GpIbβ, GpIX and filamin A. The expression of GpIbα with a C-terminal deletion between residues 560 and 570 (which inhibits filamin A binding), but not GpIbα with alanine substitutions at C-terminal residues 557 through 559 (which converts the sequence RGS to AAA but doesn’t inhibit filamin binding), results in the near-elimination of immature GpIbα. These results suggest that GpIbα binding to filamin enhances the surface expression of GpIb-IX by directing nascent protein trafficking away from a degradative pathway and towards the golgi. This leads to increased glycosylation, which further stabilizes the complex by attenuating the susceptibility of the extracellular domain to proteolytic cleavage.

scholarly journals Improving the Solubility and Digestibility of Potato Protein with an Online Ultrasound-Assisted PH Shifting Treatment at Medium Temperature

Foods ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 1908
Author(s):  
Chao Mao ◽  
Juan Wu ◽  
Xiangzhi Zhang ◽  
Fengping Ma ◽  
Yu Cheng
Keyword(s):  
Structural Changes ◽  
In Vitro Digestion ◽  
Potato Protein ◽  
Medium Temperature ◽  
Tertiary Structures ◽  
Force Microscopy ◽  
Atomic Force ◽  
Soluble Aggregates ◽  
Ph Shifting

Ultrasonic (US) treatment was combined with pH shifting (pHS) and mild thermal (40 °C) (T40) treatment (US/T40/pHS) to improve the solubility of potato protein. The effects of the ultrasonication frequency, ultrasonication time, and incorporation sequence on the solubility of potato protein were investigated. The results showed that online US/T40/pHS treatment resulted in higher solubility of potato protein and enhanced free amino group release during in vitro digestion. The solubility of potato protein treated with online US/T40/pHS at a mono-frequency of 40 kHz for 15 min increased by 1.73 times compared with the control (p < 0.05). The digestibility rate increased by 16.0% and 30.8% during gastric and intestinal digestion, respectively, compared with the control (p < 0.05). It was demonstrated that online US/T40/pHS treatment significantly changed the secondary and tertiary structures of potato protein according to the results of circular dichroism and internal fluorescence. SDS-PAGE, particle size, and atomic force microscopy (AFM) showed that structural changes led to the formation of large soluble aggregates. The results suggested that the improvement in the solubility and digestibility of potato protein treated with online US/T40/pHS may be due to the formation of large soluble aggregates, which are more hydrophilic and sensitive to digestive enzymes.

scholarly journals Interactions Under Crowding Milieu: Chemical-Induced Denaturation of Myoglobin is Determined by the Extent of Heme Dissociation on Interaction with Crowders

Biomolecules ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 490 ◽  
Author(s):  
Khalida Nasreen ◽  
Zahoor Ahmad Parray ◽  
Shahzaib Ahamad ◽  
Faizan Ahmad ◽  
Anwar Ahmed ◽  
...  
Keyword(s):  
Structural Changes ◽  
Ph Dependence ◽  
Docking Studies ◽  
Titration Calorimetry ◽  
Dextran 70 ◽  
The Stability ◽  
Single Binding Site ◽  
Intracellular Milieu

Generally, in vivo function and structural changes are studied by probing proteins in a dilute solution under in vitro conditions, which is believed to be mimicking proteins in intracellular milieu. Earlier, thermal-induced denaturation of myoglobin, in the milieu of crowder molecule showed destabilization of the metal protein. Destabilization of protein by thermal-induced denaturation involves a large extrapolation, so, the reliability is questionable. This led us to measure the effects of macromolecular crowding on its stability by chemical-induced denaturation of the protein using probes like circular dichroism and absorption spectroscopy in the presence of dextran 70 and ficoll 70 at various pHs (acidic: 6.0, almost neutral: 7.0 and basic: 8.0). Observations showed that the degree of destabilization of myoglobin was greater due to ficoll 70 as compared to that of dextran 70 so it can be understood that the nature of the crowder or the shape of the crowder has an important role towards the stability of proteins. Additionally, the degree of destabilization was observed as pH dependent, however the pH dependence is different for different crowders. Furthermore, isothermal titration calorimetry and molecular docking studies confirmed that both the crowders (ficoll and dextran) bind to heme moiety of myoglobin and a single binding site was observed for each.

Patterns of apoptosis in the corpora lutea of the rat during the oestrous cycle, pregnancy and in vitro culture

2001 ◽  
Vol 13 (3) ◽  
pp. 105 ◽  
Author(s):  
N. W. Bruce ◽  
S. Hisheh ◽  
A. M. Dharmarajan
Keyword(s):  
In Vitro Culture ◽  
Structural Changes ◽  
Molecular Mechanisms ◽  
Apoptotic Cell ◽  
Post Partum ◽  
Oestrous Cycle ◽  
Cycle Phase ◽  
Corpora Lutea ◽  
Functional Regression

Apoptosis is associated with regression of corpora lutea (CL) in a number of species. The present work compared apoptotic rates, assessed by low molecular weight DNA labelling, in rat CL of the oestrous cycle, pregnancy, the immediate post-partum period, and in vitro culture. Apoptosis was significantly related to cycle phase with peak activity at oestrus. This pattern was similar in CL formed from the most recent ovulation and from the two previous generations. Apoptosis was evident during pregnancy, albeit at low rates. It declined on the day of parturition and increased on the following day. Apoptosis was greatly increased in cultured CL to reach 20-fold higher levels than those achieved during the oestrous cycle or pregnancy. Collectively, these results suggest that apoptosis has lesser significance in rat CL functional regression than in other species with more precipitate structural changes. However, rat CL clearly retain the potential for high rates of apoptosis and so present a useful model for examining molecular mechanisms of apoptotic cell death.

Exploration of Fungal Lipase as Direct Target of Eugenol through Spectroscopic Techniques

2019 ◽  
Vol 26 (12) ◽  
pp. 919-929
Author(s):  
Farheen Naz ◽  
Haider Anis ◽  
Ziaul Hasan ◽  
Asimul Islam ◽  
Luqman A. Khan
Keyword(s):  
Bovine Serum Albumin ◽  
Serum Albumin ◽  
Binding Sites ◽  
Bovine Serum ◽  
Structural Changes ◽  
Tertiary Structure ◽  
Spectroscopic Techniques ◽  
Vis Spectroscopy ◽  
Direct Target

Background:Fungal lipase dependent processes are important for their pathogenicity. Lipases can therefore be explored as direct target of promising herbal antifungals.Objective:We explored Aspergillus niger lipase as a direct target of eugenol through spectroscopic techniques and compare results with Bovine Serum Albumin and lysozyme to comment on selectivity of eugenol towards lipase.Methods:In vitro activity assays of lipase are used to determine concentration ranges. UV-Visible, Fluorescence and Circular dichroism spectroscopy were employed to determine binding constant, stoichiometric binding sites and structural changes in Lipase, BSA and lysozyme following incubation with varying concentrations of eugenol.Results:In activity assays 50% inhibition of lipase was obtained at 0.913 mmoles/litre eugenol. UV-vis spectroscopy shows formation of lipase-eugenol, Bovine Serum Albumin-eugenol and lysozyme-eugenol complex well below this concentration of eugenol. Eugenol binding caused blue shift with Bovine Serum Albumin and lysozyme suggestive of compaction, and red shift with lipase. Negative ellipticity decreased with lipase but increased with Bovine Serum Albumineugenol and lysozyme-eugenol complexes suggesting loss of helical structure for lipase and compaction for Bovine Serum Albumin and lysozyme. Binding of eugenol to lipase was strong (Ka= 4.7 x 106 M-1) as compared to Bovine Serum Albumin and lysozyme. The number of stoichiometric eugenol binding sites on lipase was found to be 2 as compared to 1.37 (Bovine Serum Albumin) and 0.32 (lysozyme). Docking results also suggest strong binding of eugenol with lipase followed by Bovine Serum Albumin and lysozyme.Conclusion:Eugenol is found to be effective inhibitor and disruptor of secondary and tertiary structure of lipase, whereas its binding to Bovine Serum Albumin and lysozyme is found to be weak and less disruptive of structures suggesting selectivity of eugenol towards lipase.

scholarly journals Conventional frequency ultrasound detection of tumor response in vivo to cancer treatment administration

2021 ◽  
Author(s):  
Naum Papanicolau
Keyword(s):  
Cell Death ◽  
Cancer Therapy ◽  
Structural Changes ◽  
Apoptotic Cell ◽  
High Frequency Ultrasound ◽  
Patient Response ◽  
Tumor Morphology ◽  
Frequency Ultrasound

Current methods employed to evaluate patient response to cancer therapy are typically invasive requiring examination of excised tissue. The development of a non-invasive method of monitoring patient response to cancer therapy administration would potentiate clinical decisions permitting clinicians to adjust therapy regimens early in a treatment course based upon individual patient responses. It has been previously demonstrated that high frequency ultrasound is capable of reliably quantifying structural changes in tumor morphology in response to cancer therapies. Preliminary work has also indicated that ultrasound employed at clinically relevant frequencies (1-15 MHz) can detect apoptotic cell death using in vitro models. This thesis examines changes in tumor morphology in response to cancer therapy administration employing ultrasound at a clinically applicable frequency in a preclinical in vivo mouse model. The power spectrum of the radiofrequency data obtained from tumors was analyzed via linear regression spectroscopic analysis, as well as evaluating a statistical analysis of the amplitude distribution of the signal envelope. It is demonstrated here for the first time that 7 MHz ultrasound can detect apoptotic and other forms of cell death in vivo. A potential for a parametric imaging technique to visually represent analysis results is also demonstrated.

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