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1.
Rashi Verma Monika Yadav Rajabrata Bhuyan Shweta Aggarwal Arnab Nayek 《Journal of receptor and signal transduction research》2016,36(6):601-616
Computer-aided antibody engineering has been successful in the design of new biologics for disease diagnosis and therapeutic interventions. Interleukin-6 (IL-6), a well-recognized drug target for various autoimmune and inflammatory diseases such as rheumatoid arthritis, multiple sclerosis, and psoriasis, was investigated in silico to design potential lead antibodies. Here, crystal structure of IL-6 along with monoclonal antibody olokizumab was explored to predict antigen–antibody (Ag???Ab)-interacting residues using DiscoTope, Paratome, and PyMOL. Tyr56, Tyr103 in heavy chain and Gly30, Ile31 in light chain of olokizumab were mutated with residues Ser, Thr, Tyr, Trp, and Phe. A set of 899 mutant macromolecules were designed, and binding affinity of these macromolecules to IL-6 was evaluated through Ag???Ab docking (ZDOCK, ClusPro, and Rosetta server), binding free-energy calculations using Molecular Mechanics/Poisson Boltzman Surface Area (MM/PBSA) method, and interaction energy estimation. In comparison to olokizumab, eight newly designed theoretical antibodies demonstrated better result in all assessments. Therefore, these newly designed macromolecules were proposed as potential lead antibodies to serve as a therapeutics option for IL-6-mediated diseases. 相似文献
2.
Mammary epithelial cells from virgin Balb/c mice were isolated by collagenase digestion and cultured within collagen gels in serum-free basal medium containing insulin (10 micrograms/ml). Previous work has shown that linoleate or its metabolite, prostaglandin E2 (PGE2), stimulate the growth of these cells only in the presence of a growth stimulant such as epidermal growth factor (EGF). Since PGE2 can stimulate cyclic AMP (cAMP) production, the role of cAMP in linoleate and EGF-stimulated growth was examined. The cAMP phosphodiesterase inhibitor, IBMX (0.1 mM), was found to augment growth when cells were cultured in the presence of both EGF and linoleate or PGE2, but not either factor alone. These results indicated that EGF does not stimulate proliferation via cyclic AMP mediated events but could synergize with cAMP events if cAMP levels were elevated by PGE2. When assayed in cells plated on top of collagen-coated culture dishes, cellular cyclic AMP levels were stimulated by PGE2, but only marginally by EGF. Although the stimulation of endogenous cAMP by PGE2 and IBMX was insufficient to stimulate growth in the absence of EGF, exogenous dibutyryl-cAMP (greater than 100 micrograms/ml) was able to do so showing that a sustained, and high level of cAMP (greater than 100 micrograms/ml) could stimulate growth in insulin-containing basal medium. EGF was capable of enhancing the cellular sensitivity to dibutyryl-cAMP but the converse was not observed. cAMP stimulation of growth was dependent upon a superphysiological concentration of insulin (10 micrograms/ml) or a physiological concentration of somatomedin-C. These results indicate that the proliferation of mouse mammary epithelial cells can be stimulated separately or in synergism by cAMP-dependent or -independent events. 相似文献
3.
Efficient selection intensity in early generation index selection in groundnut (Arachis hypogaea L.)
A. Bandyopadhyay V. Arunachalam K. Venkaiah 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1985,71(2):300-304
Summary The F2 potential of single and three-way crosses was evaluated using a set of physiological and yield components. Results were based on an index of selection using (a) only yield components and (b) both physiological and yield components. The indices were constructed using the percentage improvement of F2 over the better parent of the corresponding F1 cross for every character. The performance of F2 plants assessed by the expected value of the regression index was ranked in descending order to provide a ranked F2 distribution (FRD). The FRD was divided into four equal parts, T25 (top 25%), T50 (26–50%), T75 (51–75%) and T100 (76–100%). F3 families derived from F2 plants in T25 were found to provide a higher frequency of selections for pod number than T50, T75 and T100. The frequency of selections was higher in three-way than single crosses. Selection index based on physiological and yield components was more efficient in trapping F2 plants providing selections in F3 than the index based on yield components only. The results brought out the importance of bunch x bunch crosses as a complement to the usually advocated bunch x runner ones. 相似文献
4.
Controlling certain diseases using peptide drugs has remarkably increased in the past two decades. In this regard, a generic formulation is an upfront solution to fulfill market demands. Ganirelix, a leading peptide active pharmaceutical ingredient (API) primarily used as a gonadotropin-releasing hormone antagonist (GnRH), has established a potential market value worldwide. But its generic formulation mandates detailed impurity profiles from a synthetic source and contemplates the sameness of a reference-listed drug (RLD). Post-chemical synthesis and processing of Ganirelix, some commercial sources have revealed two new potential impurities among many known, which show the deletion of an ethyl group from the hArg(Et)2 residue at the sixth and eighth positions, named des-ethyl-Ganirelix. These impurities are unprecedented in traditional peptide chemistry, and such monoethylated-hArg building blocks are not easily accessible commercially to synthesize these two impurities. Here, we have outlined the synthesis, purification, and enantiomeric purity characterization of the amino acids and their incorporation in the Ganirelix peptide sequence to synthesize these potential peptide impurities. This methodology will enable the convenient synthesis of side-chain substituted Arg and hArg derivatives in peptide drug discovery platforms. 相似文献
5.
We develop a new method for variable selection in a nonlinear additive function-on-scalar regression (FOSR) model. Existing methods for variable selection in FOSR have focused on the linear effects of scalar predictors, which can be a restrictive assumption in the presence of multiple continuously measured covariates. We propose a computationally efficient approach for variable selection in existing linear FOSR using functional principal component scores of the functional response and extend this framework to a nonlinear additive function-on-scalar model. The proposed method provides a unified and flexible framework for variable selection in FOSR, allowing nonlinear effects of the covariates. Numerical analysis using simulation study illustrates the advantages of the proposed method over existing variable selection methods in FOSR even when the underlying covariate effects are all linear. The proposed procedure is demonstrated on accelerometer data from the 2003–2004 cohorts of the National Health and Nutrition Examination Survey (NHANES) in understanding the association between diurnal patterns of physical activity and demographic, lifestyle, and health characteristics of the participants. 相似文献
6.
Keya Bandyopadhyay Sudipan Karmakar Aruna Biswas Pijush K Das 《European journal of biochemistry》2003,270(18):3806-3813
Earlier we presented several lines of evidence that a 67-kDa laminin binding protein (LBP) in Leishmania donovani, that is different from the putative mammalian 67-kDa laminin receptor, may play an important role in the onset of leishmaniasis, as these parasites invade macrophages in various organs after migrating through the extracellular matrix. Here we describe the membrane orientation of this Leishmania laminin receptor. Flow cytometric analysis using anti-LBP Ig revealed its surface localization, which was further confirmed by enzymatic radiolabeling of Leishmania surface proteins, autoradiography and Western blotting. Efficient incorporation of LBP into artificial lipid bilayer, as well as its presence in the detergent phase after Triton X-114 membrane extraction, suggests that it may be an integral membrane protein. Limited trypsinization of intact parasite and subsequent immunoblotting of trypsin released material using laminin as primary probe revealed that a major part of this protein harbouring the laminin binding site is oriented extracellularly. Carboxypeptidase Y treatment of the whole cell, as well as the membrane preparation, revealed that a small part of the C-terminal is located in the cytosol. A 34-kDa transmembrane part of LBP could be identified using the photoactive probe, 3-(trifluoromethyl)-3-(m-iodophenyl)diazirine (TID). Partial sequence comparison of the intact protein to that with the trypsin-released fragment indicated that N-terminal may be located extracellularly. Together, these results suggest that LBP may be an integral membrane protein, having significant portion of N-terminal end as well as the laminin binding site oriented extracellularly, a membrane spanning domain and a C-terminal cytosolic end. 相似文献
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9.
Quaternary structure and geminate recombination in hemoglobin: flow-flash studies on alpha 2CO beta 2 and alpha 2 beta 2CO.
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The kinetics of geminate recombination for the diliganded species alpha 2CO beta 2 and alpha 2 beta 2CO of human hemoglobin were studied using flash photolysis. The unstable diliganded species were generated just before photolysis by chemical reduction in a continuous flow reactor from the more stable valency hybrids alpha 2CO beta 2+ and alpha 2+ beta 2CO, which could be prepared by high pressure liquid chromatography. Before the flash photolysis studies, the hybrids had been characterized by double-mixing stopped-flow kinetics experiments. At pH 6.0 in the presence of inositol hexaphosphate (IHP) both of the diliganded species show second order kinetics for overall addition of a third CO that is clearly characteristic of the T state (l' = 1-2 x 10(5) M-1 s-1), whereas at higher pH and in the absence of IHP they show combination rates characteristic of an R state. The kinetics of geminate recombination following photolysis of a bound CO, however, showed little dependence on pH and IHP concentration. This surprising observation is explained on the basis that the kinetics of geminate recombination of CO primarily depends on the tertiary structure of the ligand binding site, which apparently does not differ much between the R state and the liganded T state formed on adding IHP in this system. Since this explanation requires distinguishing different tertiary structures within a particular quaternary structure, it amounts to a contradiction to the two-state allosteric model. 相似文献
10.
Possible involvement of protein phosphorylation in interferon (IFN)-mediated activation of IFN-stimulated gene factor 3 (ISGF3) was investigated. For this purpose, in vivo experiments with specific inhibitors of protein kinases and in vitro experiments with protein phosphatases were carried out. In HeLaM cells, 2-aminopurine, an inhibitor of double-stranded RNA-dependent protein kinase, blocked the induction of ISGF3 gamma subunit but not the activation of ISGF3 alpha subunit. A series of experiments using combinations of protein and RNA synthesis inhibitors and 2-aminopurine indicated that the block elicited by 2-aminopurine was at the level of ISGF3 gamma mRNA synthesis. Activation of ISGF3 alpha, although insensitive to 2-aminopurine, was completely blocked by 10 nM staurosporine, an inhibitor of protein kinase C. On the other hand, even 500 nM staurosporine did not block the induction of ISGF3 gamma. Incubation of cytoplasmic or nuclear extracts of IFN-treated HeLaM cells in vitro with alkaline phosphatase completely eliminated their ability to form the ISGF3 complex but not the ISGF1 complex. Treatment with acid phosphatase, on the other hand, changed the electrophoretic mobility of the ISGF3 complex but did not obliterate it. Complementation experiments revealed that ISGF3 alpha was the alkaline phosphatase-sensitive component of the complex. These results suggest that a protein kinase C-mediated phosphorylation step is involved in ISGF3 alpha activation and a 2-aminopurine-sensitive component is involved in ISGF3 gamma mRNA induction. 相似文献