Studies in the metabolism of cobalt

This thesis, after a brief review of the literature dealing with the effects of cobalt deficiency in animals and especially the physiological rôle of cobalt describes experiments designed to clarify certain aspects of these problems.The conclusions arrived at from the present investigation are as follows: -(a) That lambs grazing cobalt dressed pasture eat more grass than lambs grazing cobalt deficient pasture of equal nutritive value as far as proteins, fats and carbohydrates are concerned. This was established by grazing experiments with groups of lambs equipped with a suitable harness which allowed of the daily measurement of faeces excreted.(b) That the difference in the amount of nutrients eaten by the two groups would suggest that the clinical symptoms of 'Pining' - the diseased condition associated with cobalt deficiency - were those of malnutrition.(c) That the digestion of crude fibre by sheep on cobalt deficient diets is of a low order and can be increased by the oral administration of cobalt salts.(d) That the digestion of the other organic nutrients of the ration is not materially altered by a low cobalt intake.(e) That a daily diet containing 0.02 mg. of cobalt/ 44. cobalt does not allow of the animal being in a state of positive cobalt balance but that the addition of 3 mg. of cobalt per day changes a negative cobalt balance to a positive one with the animal retaining a large proportion of the 3 mgs. of. cobalt.(f) That the major portion of cobalt ingested is excreted in the faeces and only minute amounts in the urine.(g) That all the data obtained in the present series of experiments would support the theory that the physiological function of cobalt is connected with r»mi nation

Immunohistochemical Evaluation of p63, E-Cadherin, Collagen I and III Expression in Lower Limb Wound Healing under Honey

Honey is recognized traditionally for its medicinal properties and also appreciated as a topical healing agent for infected and noninfected wounds. This study evaluates impact of honey-based occlusive dressing on nonhealing (nonresponding to conventional antibiotics) traumatic lower limb wounds (n = 34) through clinicopathological and immunohistochemical (e.g., expression of p63, E-cadherin, and Collagen I and III) evaluations to enrich the scientific validation. Clinical findings noted the nonadherence of honey dressing with remarkable chemical debridement and healing progression within 11–15 days of postintervention. Histopathologically, in comparison to preintervention biopsies, the postintervention tissues of wound peripheries demonstrated gradual normalization of epithelial and connective tissue features with significant changes in p63+ epithelial cell population, reappearance of membranous E-cadherin (P < .0001), and optimum deposition of collagen I and III (P < .0001). Thus, the present study for the first time reports the impact of honey on vital protein expressions in epithelial and connective tissues during repair of nonhealing lower limb wounds

Molecular association of glucose-6- phosphate isomerase and pyruvate kinase M2 with glyceraldehyde-3-phosphate dehydrogenase in cancer cells

Background: For a long time cancer cells are known for increased uptake of glucose and its metabolization through glycolysis. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key regulatory enzyme of this pathway and can produce ATP through oxidative level of phosphorylation. Previously, we reported that GAPDH purified from a variety of malignant tissues, but not from normal tissues, was strongly inactivated by a normal metabolite, methylglyoxal (MG).Molecular mechanism behind MG mediated GAPDH inhibition in cancer cells is not well understood. Methods: GAPDH was purified from Ehrlich ascites carcinoma (EAC) cells based on its enzymatic activity. GAPDH associated proteins in EAC cells and 3-methylcholanthrene (3MC) induced mouse tumor tissue were detected by mass spectrometry analysis and immunoprecipitation (IP) experiment, respectively. Interacting domains of GAPDH and its associated proteins were assessed by in silico molecular docking analysis. Mechanism of MG mediated GAPDH inactivation in cancer cells was evaluated by measuring enzyme activity, Circular dichroism (CD) spectroscopy, IP and mass spectrometry analyses. Result: Here, we report that GAPDH is associated with glucose-6-phosphate isomerase (GPI) and pyruvate kinase M2 (PKM2) in Ehrlich ascites carcinoma (EAC) cells and also in 3-methylcholanthrene (3MC) induced mouse tumor tissue. Molecular docking analyses suggest C-terminal domain preference for the interaction between GAPDH and GPI. However, both C and N termini of PKM2 might be interacting with the C terminal domain of GAPDH. Expression of both PKM2 and GPI is increased in 3MC induced tumor compared with the normal tissue. In presence of 1 mM MG,association of GAPDH with PKM2 or GPI is not perturbed, but the enzymatic activity of GAPDH is reduced to 26.8 ± 5 % in 3MC induced tumor and 57.8 ± 2.3 % in EAC cells. Treatment of MG to purified GAPDH complex leads to glycation at R399 residue of PKM2 only, and changes the secondary structure of the protein complex. Conclusion: PKM2 may regulate the enzymatic activity of GAPDH. Increased enzymatic activity of GAPDH in tumor cells may be attributed to its association with PKM2 and GPI. Association of GAPDH with PKM2 and GPI could be a signature for cancer cells. Glycation at R399 of PKM2 and changes in the secondary structure of GAPDH complex could be one of the mechanisms by which GAPDH activity is inhibited in tumor cells by MG

Role of Nonmuscle Myosin II in Virus-Cell Fusion

Membrane fusion is the process whereby two separate lipid bilayers merge to become one. Despite substantial progress, an integrated concept for protein–mediated membrane fusion (cellular and viral) is not yet available, and many open questions yet to be answered. Membrane fusion, the merging/intermixing of two lipid bilayers, is quite a well known process involved in a number of physiological functions e.g. fertilization, cell division, myoblast differentiation, transport of impermeant molecules into the cell (endocytosis) and out of the cell (exocytosis). Among all the cases of membrane fusion, viral entry through membrane fusion gains the special attention as viruses’ e.g. parainfluenza virus (PIV), human immunodeficiency virus (HIV), simian immunodeficiency virus (SIV), murine leukemia virus (MLV), herpes simplex virus (HSV), measles virus (MV) etc fuse directly with plasma membrane to enter inside the host cells. These viruses are the etiological agents of many biologically important diseases of man and other animals. The involvement of membrane fusion in all these events makes ‘fusion’ a sizzling issue always and force the scientists to go into its finer details. Virus-cell surface receptor interactions can elicit two types of signals; conformational changes of viral particles, and concomitant intracellular signals triggering specific cellular reactions.1 In this direction, Wang et. al. have shown evidence that cellular signal transduction pathways and associated protein kinases could be responsible in modulating retrovirus-induced cell-cell fusion.2 However, little is known about the virus-induced host cell intracellular signaling in terms of maintaining the actomyosin complex that influences the membrane fusion. Viruses in the paramyxoviridae family and others like HIV (Human immunodeficiency virus), bind to the surface proteins of the host cell and fuse with the plasma membrane to release the nucleoprotein in the cytosol directly to reach the site of their replication. If we talk about membrane fusion in terms of thermodynamics, it is energetically a non-spontaneous process and there is a very high kinetic barrier, so the process needs the catalytic help of proteins (fusion glycoproteins of all viruses and fusogens of all other kind) for using the free energy, liberated during the conformational change of the membrane proteins to draw each other together3(Fig. 1). Moreover, as membrane fusion is the critical step in the course of enveloped animal virus infection, it is logical to think of its regulation by host cell signaling. Keeping this proposition in mind we have studied a precise effect of membrane fusion on actomyosin signaling in the course of viral infection, taking Sendai virus (Z strain) as the model.Research was conducted under the supervision of Prof. S S Jana of Biological Chemistry division under the SBS [School of Biological Sciences]The research was carried out under CSIR, DBT & DST gran

Mathematical Characterization of Protein Sequences Using Patterns as Chemical Group Combinations of Amino Acids

<div><p>Comparison of amino acid sequence similarity is the fundamental concept behind the protein phylogenetic tree formation. By virtue of this method, we can explain the evolutionary relationships, but further explanations are not possible unless sequences are studied through the chemical nature of individual amino acids. Here we develop a new methodology to characterize the protein sequences on the basis of the chemical nature of the amino acids. We design various algorithms for studying the variation of chemical group transitions and various chemical group combinations as patterns in the protein sequences. The amino acid sequence of conventional myosin II head domain of 14 family members are taken to illustrate this new approach. We find two blocks of maximum length 6 aa as ‘FPKATD’ and ‘Y/FTNEKL’ without repeating the same chemical nature and one block of maximum length 20 aa with the repetition of chemical nature which are common among all 14 members. We also check commonality with another motor protein sub-family kinesin, KIF1A. Based on our analysis we find a common block of length 8 aa both in myosin II and KIF1A. This motif is located in the neck linker region which could be responsible for the generation of mechanical force, enabling us to find the unique blocks which remain chemically conserved across the family. We also validate our methodology with different protein families such as MYOI, Myosin light chain kinase (MLCK) and Rho-associated protein kinase (ROCK), Na⁺/K⁺-ATPase and Ca²⁺-ATPase. Altogether, our studies provide a new methodology for investigating the conserved amino acids’ pattern in different proteins.</p></div

Distinct ranges of every branch of the phylogenetic tree obtained from Table 11, percentage (%) identity of each branching point from Fig 2.

<p>Distinct ranges of every branch of the phylogenetic tree obtained from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0167651#pone.0167651.t011" target="_blank">Table 11</a>, percentage (%) identity of each branching point from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0167651#pone.0167651.g002" target="_blank">Fig 2</a>.</p

Amino acid categorization based on their chemical nature.

<p>Amino acid categorization based on their chemical nature.</p

Common motifs Myosin II and KIF 1A class of Kinesin, their position and sequences.

<p>Common motifs Myosin II and KIF 1A class of Kinesin, their position and sequences.</p

Details of Rho-associated protein kinase family members in human.

<p>Details of Rho-associated protein kinase family members in human.</p

Details of MLCK protein family members in human.

<p>Details of MLCK protein family members in human.</p