Effect of pH on Diclofenac-Lysozyme Interaction: Structural and Functional Aspect.

As a nonsteroidal antiinflammatory drug, diclofenac (DCF) is used in the treatment of a variety of human ailments. It has already been reported that the use of this class of drugs for a longer duration is associated with numerous side effects such as cardiovascular implications, reno-medullary complications, etc. In the present study, the effect of DCF on the structure, stability, and function of lysozyme was studied. The study was designed to examine the effect of DCF only at various pH values. Heat-induced denaturation of lysozyme was analyzed in the presence and absence of various molar concentrations of DCF at different pH values. The values of thermodynamic parameters, the midpoint of denaturation (T m), enthalpy change at T m (ΔH m), constant pressure heat capacity change (ΔC p), and Gibbs energy change at 25°C (ΔG D o), thus obtained under a given set of conditions (pH and molar concentration of DCF), demonstrated the following 1) DCF destabilized lysozyme with respect of T m and ΔG D o at all the pH values, 2) the magnitude of protein destabilization is lesser at acidic pH than at physiological pH, 3) structural changes in lysozyme are less projecting at pH 2.0 than at pH 7.0, and 4) quenching is observed at both pH values. Furthermore, the process of protein destabilization in the presence of DCF is entropically driven.

Protecting thermodynamic stability of protein: The basic paradigm against stress and unfolded protein response by osmolytes.

Organic osmolytes are known to play important role in stress protection by stabilizing macromolecules and suppressing harmful effects on functional activity. There is existence of several reports in the literature regarding their effects on structural, functional and thermodynamic aspects of many enzymes and the interaction parameters with proteins have been explored. Osmolytes are compatible with enzyme function and therefore, can be accumulated up to several millimolar concentrations. From the thermodynamic point of view, osmolyte raises mid-point of thermal denaturation (Tm) of proteins while having no significant effect on ΔGD° (free energy change at physiological condition). Unfavorable interaction with the peptide backbone due to preferential hydration is the major driving force for folding of unfolded polypeptide in presence of osmolyte. However, the thermodynamic basis of stress protection and origin of compatibility paradigm has been a debatable issue. In the present manuscript, we attempt to elaborate the origin of stress protection and compatibility paradigm of osmolytes based on the effect on thermodynamic stability of proteins. We also infer that protective effects of osmolytes on ΔGD° (of proteins) could also indicate its potential involvement in unfolded protein response and overall stress biology on macromolecular level.

Molecular basis of pathogenic parasitic infections: insights from parasite kinome.

Infectious diseases caused by numerous parasitic pathogens represent a global health conundrum. Several animal and plant pathogens are responsible for causing acute illness in humans and deadly plant infections. These pathogens have evolved a diverse array of infection strategies and survival methods within the host organism. Recent research has highlighted the role of protein kinases in the overall virulence and pathogenicity of the pathogens. Protein kinases (Pks) are a group of enzymes known to catalyse the phosphorylation of a wide variety of cellular substrates involved in different signalling cascades. They are also involved in regulating pathogen life cycle and infectivity. In this review, we attempt to address the role of parasite kinome in host infection, pathogen survival within the host tissue and thereby disease manifestation. The understanding of the parasite kinome can be a potential target for robust diagnosis and effective therapeutics.

The Extracellular Protein, Transthyretin Is an Oxidative Stress Biomarker.

The extracellular protein, transthyretin is responsible for the transport of thyroxin and retinol binding protein complex to the various parts of the body. In addition to this transport function, transthyretin has also been involved in cardiovascular malfunctions, polyneuropathy, psychological disorders, obesity and diabetes, etc. Recent developments have evidenced that transthyretin has been associated with many other biological functions that are directly or indirectly associated with the oxidative stress, the common hallmark for many human diseases. In this review, we have attempted to address that transthyretin is associated with oxidative stress and could be an important biomarker. Potential future perspectives have also been discussed.

Testing the ability of non-methylamine osmolytes present in kidney cells to counteract the deleterious effects of urea on structure, stability and function of proteins.

Human kidney cells are under constant urea stress due to its urine concentrating mechanism. It is believed that the deleterious effect of urea is counteracted by methylamine osmolytes (glycine betaine and glycerophosphocholine) present in kidney cells. A question arises: Do the stabilizing osmolytes, non-methylamines (myo-inositol, sorbitol and taurine) present in the kidney cells also counteract the deleterious effects of urea? To answer this question, we have measured structure, thermodynamic stability (ΔG D (o)) and functional activity parameters (K m and k cat) of different model proteins in the presence of various concentrations of urea and each non-methylamine osmolyte alone and in combination. We observed that (i) for each protein myo-inositol provides perfect counteraction at 1∶2 ([myo-inositol]:[urea]) ratio, (ii) any concentration of sorbitol fails to refold urea denatured proteins if it is six times less than that of urea, and (iii) taurine regulates perfect counteraction in a protein specific manner; 1.5∶2.0, 1.2∶2.0 and 1.0∶2.0 ([taurine]:[urea]) ratios for RNase-A, lysozyme and α-lactalbumin, respectively.

Why is glycine not a part of the osmoticum in the urea-rich cells?

Kidney cells of animals including human and marine invertebrates contain high amount of the protein denaturant, urea. Methylamine osmolytes are generally believed to offset the harmful effects of urea on proteins in vitro and in vivo. In this study we have investigated the possibility of glycine to counteract the effects of urea on three proteins by measuring thermodynamic stability, ΔGD° and functional activity parameters (K(m) and k(cat)). We discovered that glycine does not counteract the effects of urea in terms of both protein stability and functional activity. We also observed that the glycine alone is compatible with enzymes function and increases protein stability in terms of T(m) (midpoint of thermal denaturation) to a great extent. Our study indicates that a most probable reason for the absence of a stabilizing osmolyte, glycine in the urea-rich cells is due to the fact that this osmolyte is non-protective to macromolecules against the hostile effects of urea, and hence is not chosen by evolutionary selection pressure.