Synthesis and Characterization of Cholesteryl Conjugated Lysozyme (CHLysozyme)

Hydrophobic interaction is important for protein conformation. Conjugation of a hydrophobic group can introduce intermolecular hydrophobic contacts that can be contained within the molecule. It is possible that a strongly folded state can be formed in solution compared with the native state. In this study, we synthesized cholesteryl conjugated lysozyme (CHLysozyme) using lysozyme and cholesterol as the model protein and hydrophobic group, respectively. Cholesteryl conjugation to lysozyme was confirmed by nuclear-magnetic resonance. Differential-scanning calorimetry suggested that CHLysozyme was folded in solution. CHLysozyme secondary structure was similar to lysozyme, although circular dichroism spectra indicated differences to the tertiary structure. Fluorescence measurements revealed a significant increase in the hydrophobic surface of CHLysozyme compared with that of lysozyme; CHLysozyme self-associated by hydrophobic interaction of the conjugated cholesterol but the hydrophobic surface of CHLysozyme decreased with time. The results suggested that hydrophobic interaction changed from intramolecular interaction to an intermolecular interaction. Furthermore, the relative activity of CHLysozyme to lysozyme increased with time. Therefore, CHLysozyme likely forms a folded state with an extended durability of activity. Moreover, lysozyme was denatured in 100% DMSO but the local environment of tryptophan in CHLysozyme was similar to that of a native lysozyme. Thus, this study suggests that protein solution stability and resistance to organic solvents may be improved by conjugation of a hydrophobic group.

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Clozapine-carboxylic acid plasticized co-amorphous dispersions: Preparation, characterization and solution stability evaluation

Acta Pharmaceutica ◽

10.1515/acph-2015-0014 ◽

2015 ◽

Vol 65 (2) ◽

pp. 133-146 ◽

Cited By ~ 18

Author(s):

Ahmed Mahmoud Abdelhaleem Ali ◽

Adel Ahmed Ali ◽

Ibrahim Abdullah Maghrabi

Keyword(s):

Carboxylic Acid ◽

Tartaric Acid ◽

Molar Ratio ◽

Sublingual Administration ◽

Solution Stability ◽

X Ray Diffraction ◽

Scanning Calorimetry ◽

Great Chance ◽

Amorphous Dispersions ◽

Pass Metabolism

Abstract This study addressed the possibility of forming of co-amorphous systems between clozapine (CZ) and various carboxylic acid plasticizers (CAPs). The aim was to improve the solubility and oral bioavailability of clozapine. Co-amorphous dispersions were prepared using modified solvent evaporation methodology at drug/plasticizer stoichiometric ratios of 1:1, 1:1.5 and 1:2. Solid state characterization was performed using differential scanning calorimetry, X-ray diffraction and infra red spectroscopy. Highly soluble homogeneous co-amorphous dispersions were formed between clozapine and CAPs via hydrogen bonding. The co-amorphous dispersions formed with tartaric acid (1:2) showed the highest dissolution percentage (> 95 % in 20 minutes) compared to pure crystalline CZ (56 %). Highly stable solutions were obtained from co-amorphous CZ-citric and CZ-tartaric acid at 1:1.5 molar ratio. The prepared dispersions suggest the possibility of peroral or sublingual administration of highly soluble clozapine at a reduced dose with the great chance to bypass the first pass metabolism.

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Molten globule state of human placental cystatin (HPC) at low pH conditions and the effects of trifluoroethanol (TFE) and methanol

Biochemistry and Cell Biology ◽

10.1139/o05-171 ◽

2006 ◽

Vol 84 (2) ◽

pp. 126-134 ◽

Cited By ~ 3

Author(s):

Fouzia Rashid ◽

Sandeep Sharma ◽

M A Baig ◽

Bilqees Bano

Keyword(s):

Conformational Changes ◽

Tertiary Structure ◽

Molten Globule ◽

Structural Features ◽

Cd Spectroscopy ◽

Native State ◽

Molten Globule State ◽

Fluorescence Measurements ◽

Helical Content ◽

Human Placental Cystatin

Acid-induced conformational changes were studied in human placental cystatin (HPC) in terms of circular dichroism (CD) spectroscopy, the binding of hydrophobic dye 1-anilinonapthalene-8-sulphonic acid (ANS), and intrinsic fluorescence measurements. Our results show the formation of an acid-induced molten globule state at pH 2.0, with significant secondary and tertiary interactions that resemble the native state, exposed hydrophobic regions and the effects of trifluoroethanol (TFE) and methanol in conversion of the acid-denatured state of HPC to the alcohol-induced state, which is characterized by increased helical content, disrupted tertiary structure, and the absence of hydrophobic clusters. Alcohol-induced formation of α-helical structures at pH 2.0 is evident from the increase in the ellipticity values at 222 nm, with native-like secondary structural features at 40% TFE. The increase in helical content was observed up to 80% TFE concentration. The ability of TFE (40%) to refold acid-denatured HPC to native-state conformation is also supported by intrinsic and ANS fluorescence measurements.Key words: human placental cystatin, molten globule, acid-induced state, trifluoroethanol, methanol, CD spectroscopy, ANS fluorescence, pH, protein folding.

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Inhibitory effect of ethacrynic acid on chloride permeability

American Journal of Physiology-Legacy Content ◽

10.1152/ajplegacy.1976.231.5.1485 ◽

1976 ◽

Vol 231 (5) ◽

pp. 1485-1489 ◽

Cited By ~ 31

Author(s):

R Motais ◽

JL Cousin

Keyword(s):

Hydrophobic Interaction ◽

Ethacrynic Acid ◽

Chloride Transport ◽

Direct Proof ◽

Inhibitory Effect ◽

Ringer Solution ◽

Relative Activity ◽

Chloride Permeability ◽

Sh Group ◽

Highly Correlated

Ethacrynic acid inhibits anion movements in ox red blood cells. The I50 for chloride is 7 X 10(-6) M. The inhibitory effect is instantaneous and completely reversed by washing the cells with a Ringer solution, suggesting that reaction with a membrane SH group is not involved in this process. Direct proof that ethacrynic acid does not act by its reactivity with thiol groups is given by experiments with dihydroethacrynic acid, a derivative that lacks the ability to combine with SH groups: the characteristics of inhibition are strictly identical (instantaneous and reversible; I50 equals 9 X 10(-6) M). All the phenoxyacetic derivatives tested were also more or less inhibitory. The relative activity of all the derivatives was highly correlated with their liposolubility, indicating that hydrophobic interaction is important in determining drug effect and influence of steric factors is minimal. The data suggest that inhibition essentially results from a hydrophobic interaction between ethacrynic acid and apolar regions of the membrane protein allowing chloride transport.

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EXAFS Study of Annealing Effects on Mg65Cu25Gd10 Metallic Glass

Materials Science Forum ◽

10.4028/www.scientific.net/msf.546-549.477 ◽

2007 ◽

Vol 546-549 ◽

pp. 477-480

Author(s):

Guo Qiang Li ◽

Li Jing Zheng ◽

Z.F. Zhang ◽

Huan Xi Li ◽

M.F. Hashmi ◽

...

Keyword(s):

Structural Changes ◽

Local Environment ◽

Test Machine ◽

Compression Tests ◽

Scanning Calorimetry ◽

X Ray ◽

Heat Treated ◽

Annealing Effects ◽

Exafs Study ◽

X Ray Absorption

The effect of heat treatment on mechanical prosperities of Mg65Cu25Gd10 glass has been studied in this paper. The thermal stability and the structure of as-spun and heat treated Mg65Cu25Gd10 alloys were examined by differential scanning calorimetry (DSC) and X-ray diffractometry (XRD). The compression tests were performed using a computer controlled tensile test machine. Extended X-ray absorption fine structure (EXAFS) was applied to study the structural changes of Mg65Cu25Gd10 glass during annealing. Results showed that the decline of mechanical properties for Mg65Cu25Gd10 glass after annealing was closely related to the changes in local environment of Cu atoms and appearance of crystalline phases, and not the Gd local structure.

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COST-FP1105: Properties of PLA films reinforced with unmodified and acetylated freeze dried nanofibrillated cellulose

Holzforschung ◽

10.1515/hf-2016-0096 ◽

2016 ◽

Vol 70 (12) ◽

pp. 1125-1134 ◽

Cited By ~ 6

Author(s):

Vesna Žepič ◽

Ida Poljanšek ◽

Primož Oven ◽

Matjaž Čop

Keyword(s):

Heterogeneous System ◽

Cellulose Nanofibrils ◽

Composite Films ◽

Hydrophobic Surface ◽

Nanofibrillated Cellulose ◽

Nanocomposite Films ◽

Poly Lactic Acid ◽

Scanning Calorimetry ◽

Elongation At Break ◽

Freeze Dried

Abstract Freeze dried nanofibrils were acetylated in a heterogeneous system with acetic anhydride, pyridine, and dimethylformamide and the obtained acetylated cellulose nanofibrils (CNFac) were combined with poly(lactic acid) (PLA) to a composite. CNFac with its partially hydrophobic surface showed a good compatibility with PLA resulting in composite films with improved properties. Tensile strength (TS), modulus of elasticity (MOE), and elongation at break (EB) of PLA/CNF increased significantly when 2–5% of CNFac was added to the PLA matrix, while the addition of 10% and higher amounts CNFac decreased the EB at a higher TS and MOE. Mechanical parameters did not improve in the case of unmodified CNF addition. The addition of CNFac maintained transparency and had absorbance values between those of pure PLA film and PLA film with 2% CNF, while films formed with the addition of 5 and 10% of CNF were less transparent. The addition of CNF did not essentially affect the thermal properties of nanocomposite films. The addition of 2–10% of CNFac increased the enthalpy and maximal temperature of cold crystallization as opposed to higher loading of CNFac. The results of differential scanning calorimetry (DSC) coincide with those of the mechanical properties. Tailoring properties of PLA/CNF are only reproducible in case of homogenously distributed CNF within the PLA matrix and by an improved interphase adhesion between PLA and CNFac.

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Transition Metal Complexes of Naproxen: Synthesis, Characterization, Forced Degradation Studies, and Analytical Method Verification

Journal of Analytical Methods in Chemistry ◽

10.1155/2016/3560695 ◽

2016 ◽

Vol 2016 ◽

pp. 1-10 ◽

Cited By ~ 7

Author(s):

Md. Sharif Hasan ◽

Ruhul Kayesh ◽

Farida Begum ◽

S. M. Abdur Rahman

Keyword(s):

Transition Metal ◽

Metal Complexes ◽

Transition Metal Complexes ◽

High Performance ◽

Parent Drug ◽

Reversed Phase ◽

Hplc Method ◽

Forced Degradation ◽

Solution Stability ◽

Scanning Calorimetry

The aim of our current research was to synthesize some transition metal complexes of Naproxen, determine their physical properties, and examine their relative stability under various conditions. Characterizations of these complexes were done by 1H-NMR, Differential Scanning Calorimetry (DSC), FT-IR, HPLC, and scanning electron microscope (SEM). Complexes were subjected to acidic, basic, and aqueous hydrolysis as well as oxidation, reduction, and thermal degradation. Also the reversed phase high-performance liquid chromatography (RP-HPLC) method of Naproxen outlined in USP was verified for the Naproxen-metal complexes, with respect to accuracy, precision, solution stability, robustness, and system suitability. The melting points of the complexes were higher than that of the parent drug molecule suggesting their thermal stability. In forced degradation study, complexes were found more stable than the Naproxen itself in all conditions: acidic, basic, oxidation, and reduction media. All the HPLC verification parameters were found within the acceptable value. Therefore, it can be concluded from the study that the metal complexes of Naproxen can be more stable drug entity and offer better efficacy and longer shelf life than the parent Naproxen.

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Subpopulations of soluble, misfolded proteins commonly bypass chaperones: How it happens at the molecular level

10.1101/2021.08.18.456736 ◽

2021 ◽

Author(s):

Ritaban Halder ◽

Daniel A. Nissley ◽

Ian Sitarik ◽

Edward P. O’Brien

Keyword(s):

Tertiary Structure ◽

Molecular Level ◽

Meta Analysis ◽

Hydrophobic Surface ◽

Misfolded Proteins ◽

Similar Degree ◽

Functional Protein ◽

Folded Proteins ◽

And Function

ABSTRACTSubpopulations of soluble, misfolded proteins can bypass chaperones within cells. The scope of this phenomenon and the lifetimes of these states have not been experimentally quantified, and how such misfolding happens at the molecular level is poorly understood. We address the first issue through a meta-analysis of the experimental literature. We find that in all quantitative protein refolding-function studies, there is always a subpopulation of soluble but misfolded and less-functional protein that does not fold in the presence of one or more chaperones. This subpopulation ranges from 8% to 50% of the soluble protein molecules in solution. Fitting the experimental time traces to a kinetic model, we find these chaperone-bypassing misfolded states take months or longer to fold and function in the presence of different chaperones. We next addressed how, at the molecular level, some misfolded proteins can evade chaperones by simulating six different proteins interacting with E. coli’s GroEL and HtpG chaperones when those proteins are in folded, unfolded, or long-lived, soluble, misfolded states. We observe that both chaperones strongly bind the unfolded state and weakly bind the folded and misfolded states to a similar degree. Thus, these chaperones cannot distinguish between the folded and long-lived misfolded states of these proteins. A structural analysis reveals the misfolded states are highly similar to the native state – having a similar size, amount of exposed hydrophobic surface area, and level of tertiary structure formation. These results demonstrate that in vitro it is common for appreciable subpopulations of proteins to remain misfolded, soluble, and evade the refolding action of chaperones for very long times. Further, these results suggest that this happens because these misfolded subpopulations are near-native and therefore interact with chaperones to a similar extent as properly folded proteins. More broadly, these results indicate a mechanism in which long-time scale changes in protein structure and function can persist in cells because some protein’s non-native states can bypass components of the proteostasis machinery.TEASERNear-native, misfolded protein conformations explain why some soluble proteins fail to refold in the presence of chaperones.

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Directed Evolution of Thermus Maltogenic Amylase toward Enhanced Thermal Resistance

Applied and Environmental Microbiology ◽

10.1128/aem.69.8.4866-4874.2003 ◽

2003 ◽

Vol 69 (8) ◽

pp. 4866-4874 ◽

Cited By ~ 71

Author(s):

Young-Wan Kim ◽

Ji-Hye Choi ◽

Jung-Wan Kim ◽

Cheonseok Park ◽

Jung-Woo Kim ◽

...

Keyword(s):

Tertiary Structure ◽

Hydrophobic Interactions ◽

Specific Activity ◽

Dynamic Change ◽

Evolutionary Process ◽

Mutant Enzyme ◽

Dna Shuffling ◽

Wild Type ◽

Scanning Calorimetry ◽

Maltogenic Amylase

ABSTRACT The thermostability of maltogenic amylase from Thermus sp. strain IM6501 (ThMA) was improved greatly by random mutagenesis using DNA shuffling. Four rounds of DNA shuffling and subsequent recombination of the mutations produced the highly thermostable mutant enzyme ThMA-DM, which had a total of seven individual mutations. The seven amino acid substitutions in ThMA-DM were identified as R26Q, S169N, I333V, M375T, A398V, Q411L, and P453L. The optimal reaction temperature of the recombinant enzyme was 75°C, which was 15°C higher than that of wild-type ThMA, and the melting temperature, as determined by differential scanning calorimetry, was increased by 10.9°C. The half-life of ThMA-DM was 172 min at 80°C, a temperature at which wild-type ThMA was completely inactivated in less than 1 min. Six mutations that were generated during the evolutionary process did not significantly affect the specific activity of the enzyme, while the M375T mutation decreased activity to 23% of the wild-type level. The molecular interactions of the seven mutant residues that contributed to the increased thermostability of the mutant enzyme with other adjacent residues were examined by comparing the modeled tertiary structure of ThMA-DM with those of wild-type ThMA and related enzymes. The A398V and Q411L substitutions appeared to stabilize the enzyme by enhancing the interdomain hydrophobic interactions. The R26Q and P453L substitutions led potentially to the formation of genuine hydrogen bonds. M375T, which was located near the active site of ThMA, probably caused a conformational or dynamic change that enhanced thermostability but reduced the specific activity of the enzyme.

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Properties of Different Varieties of Durian

Applied Sciences ◽

10.3390/app11125653 ◽

2021 ◽

Vol 11 (12) ◽

pp. 5653

Author(s):

Supeeraya Arsa ◽

Angkana Wipatanawin ◽

Rachit Suwapanich ◽

Orachorn Makkerdchoo ◽

Niphattha Chatsuwan ◽

...

Keyword(s):

Human Serum ◽

Serum Proteins ◽

Natural Antioxidants ◽

Scanning Calorimetry ◽

Durio Zibethinus ◽

Fluorescence Measurements ◽

Novel Biomarkers ◽

Artery Disease ◽

First Time

Durian (Durio zibethinus Murr.), like many other exotic, tropical, and conventional fruits, is important in the prevention of different diseases. In this study, the characterization of the main bioactive compounds of the most popular cultivars of durian and their properties are described. The changes in the quality indices of the antioxidant status were determined by CUPRAC, ABTS, FRAP, DPPH, and ORAC assays. The profiling of phytochemicals was carried out by Fourier transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC). For the first time, in vitro studies were performed by the interaction of extracted durian polyphenols with human serum proteins (HSP) such as human serum albumin (HSAlb), fibrinogen (HSFib) and globulin (HSGlo) as novel biomarkers of coronary artery disease (CAD). The fluorescence measurements of the resulting intensity and calculated binding properties of the interaction of polyphenols with proteins showed that the most reactive was Monthong durian cultivar. This study suggests that durian cultivars have relatively strong antioxidant, binding, and health potentials and could be a significant source of natural antioxidants used in daily fresh consumption and for functional foods.

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Molecular weight, tertiary structure, water binding and colon behaviour of ispaghula husk fibre

Proceedings of The Nutrition Society ◽

10.1079/pns2002216 ◽

2003 ◽

Vol 62 (1) ◽

pp. 211-216 ◽

Cited By ~ 15

Author(s):

Saphwan Al-Assaf ◽

Glyn O. Phillips ◽

Peter A. Williams ◽

Shoji Takigami ◽

Peter Dettmar ◽

...

Keyword(s):

Molecular Weight ◽

Water Content ◽

Tertiary Structure ◽

Free Water ◽

Gel Permeation Chromatography ◽

State Matrix ◽

Scanning Calorimetry ◽

Water Binding ◽

Mechanism Of Interaction ◽

A Chain

Molecular variables, using aqueous and alkaline extracts, of the polysaccharide from ispaghula husk (IH) were examined using gel-permeation chromatography linked to multi-angle laser light scattering. Progressive extraction can yield a component with a molecular weight (MW) value up to about 7×106 Da, and gels, which accompany the extraction, have MW ranging from 10–20× 106 Da. To mimic the polysaccharide degradation, particularly in the colon, the solid IH was degraded progressively using ionising radiation. A chain break occurs every 7.5 kGy in NaOH and every 15 kGy in water. The solid-state matrix is opened by the radiation to yield increased visco-elasticity of the aqueous extracts at critical radiation doses, before further degradation occurs after about 12 kGy. Differential scanning calorimetry is used to study the mechanism of interaction of water with IH. The first water to be taken up is non-freezing water and represents about twelve water molecules/disaccharide unit of the polysaccharide. As the water content is increased, the water becomes bound to the polysaccharide and freezes and melts at a temperature different from free water. This water is thermodynamically distinguishable from free water. It forms amorphous ice on cooling which crystallises exothermically and subsequently melts endothermically. Saturation occurs at a water content of 2–3 g water/g polymer, showing that about 60% of the water in the system is ‘bound’. The most surprising conclusion is that despite the fact that the IH swells in water to form a solid and stiff gel, the greater part of that water in the gel is still free and behaves like liquid water.

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