Structural dynamics of nucleosome mediated by acetylations at H3K56 and H3K115,122.

Post translational modifications have a profound role in the regulation of several biological processes such as transcription, replication, and DNA repair. Acetylation and phosphorylation form a major class of post translational modifications involved in nucleosomal regulation by modifying its structure. The effect of post translational modifications on nucleosome structure could be better explored when the molecular trajectories explaining the time dependent structural evolution over a period of time is examined at the atomic level. The present study attempts to highlight the importance of acetylation, especially at entry-exit (Lys56) and dyad (Lys115 and Lys122) regions in regulating the nucleosome accessibility and mobility using all atom simulations. It is evident from this study that acetylation at Lys56, Lys115, and Lys122 introduces local changes in the electrostatic nature of the lateral surface and thereby weakens the histone-DNA interactions. In addition, simulations also reveal significant changes in the dynamics of superhelical DNA. The acetylation at Lys56 promotes a high amplitude out-of-planar movement of entry-exit termini. Whereas, acetylation at Lys115 and Lys122 increases the flexibility of the superhelical DNA to facilitate the rolling of the superhelical DNA around the octameric histone. In essence, the present study highlights the role of acetylation at Lys56, Lys115, and Lys122 in transcriptional regulation by promoting high amplitude dynamics of superhelical DNA for a possible unwrapping as well as mobility of nucleosome.

The midgut of the silkmoth Bombyx mori is able to recycle molecules derived from degeneration of the larval midgut epithelium.

The midgut represents the middle part of the alimentary canal and is responsible for nutrient digestion and absorption in insect larva. Despite the growing interest in this organ for different purposes, such as studies on morphogenesis and differentiation, stem cell biology, cell death processes and transport mechanisms, basic information on midgut development is still lacking for a large proportion of insect species. Undoubtedly, this lack of data could hinder the full exploitation of practical applications that involve midgut as their primary target. This may represent in particular a significant problem for Lepidoptera, an insect order that includes some of the most important species of high economic importance. With the aim of overcoming this fragmentation of knowledge, we performed a detailed morphofunctional analysis of the midgut of the silkworm, Bombyx mori, a representative model among Lepidoptera, during its development from the larval up to the adult stage, focusing attention on stem cells. Our data demonstrate stem cell proliferation and differentiation, not only in the larval midgut but also in the pupal and adult midgut epithelium. Moreover, we present evidence for a complex trophic relationship between the dying larval epithelium and the new adult one, which is established during metamorphosis. This study, besides representing the first morphological and functional characterization of the changes that occur in the midgut of a lepidopteron during the transition from the larva to the moth, provides a detailed analysis of the midgut of the adult insect, a stage that has been neglected up to now.

Structural dynamics of influenza A (H1N1) hemagglutinin protein: a comparative study of Indian (2018) isolate with its evolutionary neighbor, Californian (2009) strain.

This work highlights the structure and dynamics of two trimeric HA proteins of the H1N1 virus from different origins, the pandemic Californian (HACal) and its closest Indian neighbor (HAInd), reported in 2009 and 2018, respectively. Because of mutation, HAInd acquires new N-glycosylation and epitope binding sites along with mutations at RBD, which might trigger an altered viral-host interaction mechanism. Molecular dynamics simulations performed on HA trimers for a period of 250 ns reveal the highly dynamic nature of HACal trimers inherited by the flexibility of HA monomers. In the trimer, the dynamics of one monomer are more pronounced compared to others, and the enhanced dynamics of RBD especially gain attention as they plays a key role during fusion. Conversely, the mutant HAInd trimer effectively establishes more H-bond interactions, and accordingly, the trimer undergoes more stabilized dynamics with a relatively lower amplitude of RBD dynamics, as endorsed by the reduced RMSD, Rg, and SASA variations. The cooperative and anti-cooperative motions dissected for the subdomains of both strains also reveal a prominent anticorrelative motion of RBD against other subdomains. In agreement, the free energy landscape of stable HAInd is also characterized by a single lowest wide energy basin instead of the two minimum energy basins of the HACal trimer. In essence, the mutant HAInd acquires a highly stable conformation with novel functional features, which calls for (i) further investigation on the emerging mutation-mediated variation in viral-host binding mechanism and (ii) the need for further design of site-specific potential inhibitors to face future challenges.Communicated by Ramaswamy H. Sarma.

Pharmacophore based 3D-QSAR modeling and free energy analysis of VEGFR-2 inhibitors.

VEGFR-2, a transmembrane tyrosine kinase receptor is responsible for angiogenesis and has been an attractive target in treating cancers. The inhibition mechanism of structurally diverse urea derivatives, reported as VEGFR-2 inhibitors, was explored by pharmacophore modeling, QSAR, and molecular dynamics based free energy analysis.The pharmacophore hypothesis AADRR, resulted in a highly significant atom based 3D-QSAR model (r(2) = 0.94 and q(2) = 0.84). Binding free energy analysis of the docked complexes of highly active and inactive compounds, after 7 ns MD simulation, revealed the importance of van der Waals interaction in VEGFR-2 inhibition. The decomposition of binding free energy on a per residue basis disclosed that the residues in hinge region and hydrophobic pocket play a role in discriminating the active and inactive inhibitors. Thus, the present study proposes a pharmacophore hypothesis representing the identified interactions pattern and its further application as a template in screening databases to identify novel VEGFR-2 inhibitor scaffolds.

Evolution of Indian Influenza A (H1N1) Hemagglutinin Strains: A Comparative Analysis of the Pandemic Californian HA Strain.

The need for a vaccine/inhibitor design has become inevitable concerning the emerging epidemic and pandemic viral infections, and the recent outbreak of the influenza A (H1N1) virus is one such example. From 2009 to 2018, India faced severe fatalities due to the outbreak of the influenza A (H1N1) virus. In this study, the potential features of reported Indian H1N1 strains are analyzed in comparison with their evolutionarily closest pandemic strain, A/California/04/2009. The focus is laid on one of its surface proteins, hemagglutinin (HA), which imparts a significant role in attacking the host cell surface and its entry. The extensive analysis performed, in comparison with the A/California/04/2009 strain, revealed significant point mutations in all Indian strains reported from 2009 to 2018. Due to these mutations, all Indian strains disclosed altered features at the sequence and structural levels, which are further presumed to be associated with their functional diversity as well. The mutations observed with the 2018 HA sequence such as S91R, S181T, S200P, I312V, K319T, I419M, and E523D might improve the fitness of the virus in a new host and environment. The higher fitness and decreased sequence similarity of mutated strains may compromise therapeutic efficacy. In particular, the mutations observed commonly, such as serine-to-threonine, alanine-to-threonine, and lysine-to-glutamine at various regions, alter the physico-chemical features of receptor-binding domains, N-glycosylation, and epitope-binding sites when compared with the reference strain. Such mutations render diversity among all Indian strains, and the structural and functional characterization of these strains becomes inevitable. In this study, we observed that mutational drift results in the alteration of the receptor-binding domain, the generation of new variant N-glycosylation along with novel epitope-binding sites, and modifications at the structural level. Eventually, the pressing need to develop potentially distinct next-generation therapeutic inhibitors against the HA strains of the Indian influenza A (H1N1) virus is also highlighted here.

Structural dynamics of the RNA dependent RNA polymerase of H1N1 strain affecting humans: a bioinformatics approach.

The Influenza flu is a pandemic disease that renders the highest risk factor to the society due to its efficient ability of airborne transmission. Studies on the H1N1 strain gained significant focus, since its pandemic outbreak in 2009 and particularly the computational studies on its structural elements significantly aided in revealing their functional uniqueness. Among the 10 structural proteins of H1N1, the RNA-dependent RNA polymerase (RdRp) heterotrimeric protein complex, which is responsible for the synthesis of viral RNA (vRNA) from the negative-sense RNA genome of the virus, is the focus of the present study. This study aimed to investigate the structural dynamics of the RdRp complex with particular emphasis on the reported 17 mutations. The mutant strain is more stabilized by strong concerted residue-residue interactions at both intra- and inter- monomeric levels. In comparison, the mutant strain is structurally flexible with enhanced stabilizing interactions. The structural dynamics of RdRp are significantly governed by the dynamics of the (i) endonuclease domain of PA, (ii) RNA-entry region of PB1 and (iii) cap-binding region of PB2. Explicitly, the cap binding region of PB2 expresses (i) a concerted motion with the RNA-entry region, along with (ii) an anti-correlated motion with the endonuclease domain of the PA subunit, which further supports the stable dynamics of cap-binding towards RNA binding. These findings contribute to the understanding of the structural dynamics associated with the pandemic and mutant structures of RdRp and render a basic knowledge for further development of novel inhibitors towards influenza flu affected humans.Communicated by Ramaswamy H. Sarma.

Molecular dynamics study of TMPA mediated dissociation of Nur77-LKB1 complex.

LKB1 protein is involved in the regulation of cell polarity by phosphorylating the AMPK under energetic stress conditions. LKB1 protein is expressed in both cytoplasm and nucleus. In the nucleus, LKB1 interacts with orphan nuclear receptor protein Nur77. It is reported that the interaction of LKB1 with Nur77 is disrupted by the small molecular ligand TMPA (ethyl 2-[2,3,4-trimethoxy-6-(1-octanoyl)phenyl]acetate), such that the LKB1 is enabled to play its role in cytoplasm and further to regulate/reduce the blood glucose level. In the present study, atomistic molecular dynamics simulations are performed to understand the dissociation mechanism of Nur77-LKB1 complex. The present study reveals that TMPAs induce an open-close motion of Nur77 which further decrease the stability of Nur77-LKB1 complex. As a consequence, the interface region in LKB1-Nur77 complex is more exposed for solvation and further releases the interactions existing between Nur77 and LKB1. Altogether, this study explains the TMPAs mediated Nur77-LKB1 complex dissociation.

Insights into the RNA binding mechanism of human L1-ORF1p: a molecular dynamics study.

The recognition and binding of nucleic acids by ORF1p, an L1 retrotransposon protein, have not yet been clearly understood due to the lack of structural knowledge. The present study attempts to identify the probable single-stranded RNA binding pathway of trimeric ORF1p using computational methods like ligand mapping methodology combined with molecular dynamics simulations. Using the ligand mapping methodology, the possible RNA interacting sites on the surface of the trimeric ORF1p were identified. The crystal structure of the ORF1p timer and an RNA molecule of 29 nucleotide bases in length were used to generate the structure of the ORF1p complex based on information on predicted binding sites as well as the functional states of the CTD. The various complexes of ORF1p-RNA were generated using polyU, polyA and L1RNA sequences and were simulated for a period of 75 ns. The observed stable interaction pattern was used to propose the possible binding pathway. Based on the binding free energy for complex formation, both polyU and L1RNA complexes were identified as stable complexes, while the complex formed with polyA was the least stable one. Furthermore, the importance of the residues in the CC domain (Lys137 and Arg141), the RRM loop (Arg206, Arg210 and Arg211) and the CTD (Arg 261 and Arg262) of all three chains in stabilizing the wrapped RNA has been highlighted in this study. The presence of several electrostatic interactions including H-bond interactions increases the affinity towards RNA and hence plays a vital role in retaining the wrapped position of RNA around ORF1p. Altogether, this study presents one of the possible RNA binding pathways of ORF1p and clearly highlights the functional state of ORF1p visited during RNA binding.

Structural dynamics of wild type and mutated forms of human L1 endonuclease and insights into its sequence specific nucleic acid binding mechanism: A molecular dynamics study.

Biomolecular recognition of proteins and nucleic acids is mainly mediated by their structural features and the molecular dynamics simulations approach has been used to explore this recognition processes at the atomic level. L1-Endonuclease, an enzyme involved in L1 retrotransposition, cleaves the TA junction DNA (5'-TTTT/AA-3') and expresses high specificity for target site recognition. The present study highlights the structural features of L1-endonuclease as well as DNA responsible for such specific recognition. Especially, the importance of ßB6-B5 hairpin loop in DNA recognition has been elucidated by analyzing the dynamics of Thr192 mutated L1-endonuclease. In addition, simulations of the endonuclease complexed with DNA substrates (sequences having TA and CG junctions) revealed the specificity of L1 endonuclease towards TA junction. Molecular dynamics simulations revealed that the ßB6-B5 hairpin loop protrudes well into the minor groove of DNA having TA junction and induces DNA bending such that the width of minor groove is increased. Such endonuclease induced bending of TA junction DNA sequence positions the scissile phosphodiester bond of DNA for cleavage. The innate property of minor groove widening in TA junction than in CG junction is utilized by the ßB6-ßB5 hairpin loop of endonuclease while recognizing the DNA sequences. The present study also highlights the role of Mg2+ cation in catalysis and attempts to explore the possible target site DNA cleavage mechanism.

The conformational feasibility for the formation of reaching dimer in ASV and HIV integrase: a molecular dynamics study.

Retroviral integrases are reported to form alternate dimer assemblies like the core-core dimer and reaching dimer. The core-core dimer is stabilized predominantly by an extensive interface between two catalytic core domains. The reaching dimer is stabilized by N-terminal domains that reach to form intermolecular interfaces with the other subunit's core and C-terminal domains (CTD), as well as CTD-CTD interactions. In this study, molecular dynamics (MD), Brownian dynamics (BD) simulations, and free energy analyses, were performed to elucidate determinants for the stability of the reaching dimer forms of full-length Avian Sarcoma Virus (ASV) and Human Immunodeficiency Virus (HIV) IN, and to examine the role of the C-tails (the last ~16-18 residues at the C-termini) in their structural dynamics. The dynamics of an HIV reaching dimer derived from small angle X-ray scattering and protein crosslinking data, was compared with the dynamics of a core-core dimer model derived from combining the crystal structures of two-domain fragments. The results showed that the core domains in the ASV reaching dimer express free dynamics, whereas those in the HIV reaching dimer are highly stable. BD simulations suggest a higher rate of association for the HIV core-core dimer than the reaching dimer. The predicted stability of these dimers was therefore ranked in the following order: ASV reaching dimer < HIV reaching dimer < composite core-core dimer. Analyses of MD trajectories have suggested residues that are critical for intermolecular contacts in each reaching dimer. Tests of these predictions and insights gained from these analyses could reveal a potential pathway for the association and dissociation of full-length IN multimers.

Structural dynamics of full-length retroviral integrase: a molecular dynamics analysis.

HIV integrase catalyzes the integration between host and viral DNA and is considered as an interesting target for treating HIV. Knowledge of the complete structure of integrase is inevitable to describe the communicative inter-domain interactions affecting the HIV integration and disintegration process and hence the study on full-length integrase turns out to be an essential task. In this investigation, a structure of full-length integrase is designed to analyze the global dynamics of integrase dimer and monomers (with and without the C-terminal, 270-288 amino acids) for a period of 20 ns. The molecular dynamics analysis and the subsequent DynDom analysis reveal (i) a stable dynamics of dimeric CCD and NTD domains and (ii) CCD-α11-mediated rotational-cum-translational CTD motion as the functional dynamics of IN dimer. This observation supports that (i) aggregation enhances the integrase activity and (ii) flexible CTD for its cis and trans coordination with CCD. The role of C-loop over the dynamics of integrase is also explored, which unveils that the spatial arrangement of integrase domains is changed during dynamics in the absence of C-loop. In essence, here we report a C-loop-dependent structural dynamics of integrase and the active dynamics of integrase in dimer. Further studies on C-loop sensing mechanism and the multimerization of integrase would provide insight into HIV integration and disintegration processes. Supplementary material. Movies generated from molecular dynamics trajectory showing the CTD dynamics of IN structures (monomers with & without C-loop and dimer) are linked online to this article. The remaining supplementary data can be downloaded from the author's server at the URL http://ramutha.bicpu.edu.in .