Variable selection in nonlinear function-on-scalar regression

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.     

Evolutionary link between the mycobacterial plasmid pAL5000 replication protein RepB and the extracytoplasmic function family of σ factors

Mycobacterial plasmid pAL5000 represents a family of plasmids found mostly in the Actinobacteria. It replicates using two plasmid-encoded proteins, RepA and RepB. While BLAST searches indicate that RepA is a replicase family protein, the evolutionary connection of RepB cannot be established, as no significant homologous partner (E < 10(-3)) outside the RepB family can be identified. To obtain insight into the structure-function and evolutionary connections of RepB, an investigation was undertaken using homology modeling, phylogenetic, and mutational analysis methods. The results indicate that although they are synthesized from the same operon, the phylogenetic affinities of RepA and RepB differ. Thus, the operon may have evolved through random breaking and joining events. Homology modeling predicted the presence of a three-helical helix-turn-helix domain characteristic of region 4 of extracytoplasmic function (ECF) σ factors in the C-terminal region of RepB. At the N-terminal region, there is a helical stretch, which may be distantly related to region 3 of σ factors. Mutational analysis identified two arginines indispensable for RepB activity, one each located within the C- and N-terminal conserved regions. Apart from analyzing the domain organization of the protein, the significance of the presence of a highly conserved A/T-rich element within the RepB binding site was investigated. Mutational analysis revealed that although this motif does not bind RepB, its integrity is important for efficient DNA-protein interactions and replication to occur. The present investigation unravels the possibility that RepB-like proteins and their binding sites represent ancient DNA-protein interaction modules.     

Sulfated homologues of heparin inhibit hepatitis C virus entry into mammalian cells

The mechanism of entry of hepatitis C virus (HCV) through interactions between the envelope glycoproteins and specific cell surface receptors remains unclear at this time. We have previously shown with the vesicular stomatitis virus (VSV)/HCV pseudotype model that the hypervariable region 1 of the HCV E2 envelope glycoprotein helps in binding with glycosaminoglycans present on the cell surface. In this study, we have examined the binding of HCV envelope glycoproteins with chemically modified derivatives of heparin. Furthermore, we have determined the functional relevance of the interaction of heparin derivatives with HCV envelope glycoproteins for infectivity by using a human immunodeficiency virus (HIV)/HCV pseudotype, a VSV/HCV pseudotype, and cell culture-grown HCV genotype 1a. Taken together, our results suggest that the HCV envelope glycoproteins rely upon O-sulfated esters of a heparin homologue to facilitate entry into mammalian cells.     

Probing binding mechanism of interleukin-6 and olokizumab: in silico design of potential lead antibodies for autoimmune and inflammatory diseases

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.     

Nitric oxide blocks cellular heme insertion into a broad range of heme proteins

Although the insertion of heme into proteins enables their function in bioenergetics, metabolism, and signaling, the mechanisms and regulation of this process are not fully understood. We developed a means to study cellular heme insertion into apo-protein targets over a 3-h period and then investigated how nitric oxide (NO) released from a chemical donor (NOC-18) might influence heme (protoporphyrin IX) insertion into seven targets that present a range of protein structures, heme ligation states, and functions (three NO synthases, two cytochrome P450's, catalase, and hemoglobin). NO blocked cellular heme insertion into all seven apo-protein targets. The inhibition occurred at relatively low (nM/min) fluxes of NO, was reversible, and did not involve changes in intracellular heme levels, activation of guanylate cyclase, or inhibition of mitochondrial ATP production. These aspects and the range of protein targets suggest that NO can act as a global inhibitor of heme insertion, possibly by inhibiting a common step in the process.     

Tunable luminescence of a synthesized furophenanthraquinone derivative: interactions with different solvents

The synthesis is described of a luminescent furophenanthraquinone derivative, 9‐methoxyphenanthro[4,3‐b]furan‐4,5‐dione (MPFD). The biological importance of tetracyclic furophenanthraquinones was considered and the tunable luminescence of MPFD in different solvents was studied to explore the nature of the specific interactions between MPFD and solvents. Observation of dual emission bands and identical nature of the fluorescence excitation spectra of MPFD monitored at the emission wavelength in polar solvents indicated the formation of two different types of species in the excited state, probably due to proton transfer from the solvent to MPFD. Luminescence intensity due to anionic species was found to be increased and the corresponding peak was red shifted with increase in the proton‐donating ability of the solvents, acting as an acid with respect to MPFD. Availability of more acidic protons in the solvent facilitated this phenomenon occurring in the excited state. MPFD also interacted with halogen‐containing solvents by forming electron donor–acceptor charge transfer (CT) complexes. This CT complex formation was dependent on the number of chlorine atoms; the position of the corresponding luminescence band varied with the polarity of the solvent. Extent of the CT increased with increase in the number of chlorine atoms in the dichloro, trichloro and tetrachloro solvents, whereas the luminescence peak due to the CT complex was found to be blue shifted with decrease in solvent polarity. Interaction of the synthesized bioactive MPFD with different solvents deserves biological importance as proton transfer and CT play pivotal roles in biology.     

TGFβ mediated LINC00273 upregulation sponges mir200a-3p and promotes invasion and metastasis by activating ZEB1

Transforming growth factor β (TGFβ) is a prominent cytokine that promotes tumor progression by activating epithelial-to-mesenchymal transition (EMT). This study indicated that TGFβ exerted metastasis by inducing zinc finger E-box binding homeobox 1 (ZEB1) and a long noncoding RNA, LINC00273, expressions in A549 cells. Knocking down LINC00273 diminished TGFβ induced ZEB1 expression as well as metastasis. Mechanistically, LINC00273 acted as a molecular sponge of microRNA (miR)-200a-3p which liberate ZEB1 to perform its prometastatic functions. LINC00273 knockdown and miR200a3p mimic transfection of A549 cells were used for validating the link between TGFβ and LINC00273 induced metastasis. RNA pulldown and luciferase assay were performed to establish mir200a-3p-LINC00273 interaction. High expressions of LINC00273, TGFβ, and ZEB1 with concurrent low miR200a-3p expression had been verified in vivo and in patient samples. Overall, LINC00273 promoted TGFβ-induced lung cancer EMT through miR-200a-3p/ZEB1 feedback loop and may serve as a potential target for therapeutic intervention in lung cancer metastasis.     

Characterization of haemoglobin from Actinorhizal plants – An in silico approach

Plant haemoglobins (Hbs), found in both symbiotic and non-symbiotic plants, are heme proteins and members of the globin superfamily. Hb genes of actinorhizal Fagales mostly belong to the non-symbiotic type of haemoglobin; however, along with the non-symbiotic Hb, Casuarina sp. posses a symbiotic one (symCgHb), which is expressed specifically in infected cells of nodules. A thorough sequence analysis of 26 plant Hb proteins, currently available in public domain, revealed a consensus motif of 29 amino acids. This motif is present in all the members of symbiotic class II Hbs including symCgHb and non-symbiotic Class II Hbs, but is totally absent in Class I symbiotic and non-symbiotic Hbs. Further, we constructed 3D structures of Hb proteins from Alnus and Casuarina through homology modelling and peeped into their structural properties. Structure-based studies revealed that the Casuarina symbiotic haemoglobin protein shows distinct stereochemical properties from that of the other Casuarina and Alnus Hb proteins. It also showed considerable structural similarities with leghemoglobin structure from yellow lupin (pdb id 1GDI). Therefore, sequence and structure analyses point to the fact that symCgHb protein shows significant resemblance to symbiotic haemoglobin found in legumes and may thus eventually play a similar role in shielding the nitrogenase from oxygen as seen in the case of leghemoglobin.     

Apigenin shows synergistic anticancer activity with curcumin by binding at different sites of tubulin

Apigenin, a natural flavone, present in many plants sources, induced apoptosis and cell death in lung epithelium cancer (A549) cells with an IC₅₀ value of 93.7 ± 3.7 μM for 48 h treatment. Target identification investigations using A549 cells and also in cell-free system demonstrated that apigenin depolymerized microtubules and inhibited reassembly of cold depolymerized microtubules of A549 cells. Again apigenin inhibited polymerization of purified tubulin with an IC₅₀ value of 79.8 ± 2.4 μM. It bounds to tubulin in cell-free system and quenched the intrinsic fluorescence of tubulin in a concentration- and time-dependent manner. The interaction was temperature-dependent and kinetics of binding was biphasic in nature with binding rate constants of 11.5 × 10⁻⁷ M⁻¹ s⁻¹ and 4.0 × 10⁻⁹ M⁻¹ s⁻¹ for fast and slow phases at 37 °C, respectively. The stoichiometry of tubulin–apigenin binding was 1:1 and binding the binding constant (K_d) was 6.08 ± 0.096 μM. Interestingly, apigenin showed synergistic anti-cancer effect with another natural anti-tubulin agent curcumin. Apigenin and curcumin synergistically induced cell death and apoptosis and also blocked cell cycle progression at G₂/M phase of A549 cells. The synergistic activity of apigenin and curcumin was also apparent from their strong depolymerizing effects on interphase microtubules and inhibitory effect of reassembly of cold depolymerized microtubules when used in combinations, indicating that these ligands bind to tubulin at different sites. In silico modeling suggested apigenin bounds at the interphase of α–β-subunit of tubulin. The binding site is 19 Å in distance from the previously predicted curcumin binding site. Binding studies with purified protein also showed both apigenin and curcumin can simultaneously bind to purified tubulin. Understanding the mechanism of synergistic effect of apigenin and curcumin could be helped to develop anti-cancer combination drugs from cheap and readily available nutraceuticals.