Quaternary association in beta-prism I fold plant lectins: Insights from X-ray crystallography, modelling and molecular dynamics

Dimeric banana lectin and calsepa, tetrameric artocarpin and octameric heltuba are mannose-specific beta-prism I fold lectins of nearly the same tertiary structure. MD simulations on individual subunits and the oligomers provide insights into the changes in the structure brought about in the protomers on oligomerization, including swapping of the N-terminal stretch in one instance. The regions that undergo changes also tend to exhibit dynamic flexibility during MD simulations. The internal symmetries of individual oligomers are substantially retained during the calculations. Energy minimization and simulations were also carried out on models using all possible oligomers by employing the four different protomers. The unique dimerization pattern observed in calsepa could be traced to unique substitutions in a peptide stretch involved in dimerization. The impossibility of a specific mode of oligomerization involving a particular protomer is often expressed in terms of unacceptable steric contacts or dissociation of the oligomer during simulations. The calculations also led to a rationale for the observation of a heltuba tetramer in solution although the lectin exists as an octamer in the crystal, in addition to providing insights into relations among evolution, oligomerization and ligand binding.

The Structure of the Thioredoxin-Triosephosphate Isomerase Complex Provides Insights into the Reversible Glutathione-Mediated Regulation of Triosephosphate Isomerase

Protein-protein interactions are crucial for many biological functions. The redox interactome encompasses numerous weak transient interactions in which thioredoxin plays a central role. Proteomic studies have shown that thioredoxin binds to numerous proteins belonging to various cellular processes, including energy metabolism. Thioredoxin has cross talk with other redox mechanisms involving glutathionylation and has functional overlap with glutaredoxin in deglutathionylation reactions. In this study, we have explored the structural and biochemical interactions of thioredoxin with the glycolytic enzyme, triosephosphate isomerase. Nuclear magnetic resonance chemical shift mapping methods and molecular dynamics-based docking have been applied in deriving a structural model of the thioredoxin-triosephosphate isomerase complex. The spatial proximity of active site cysteine residues of thioredoxin to reactive thiol groups on triosephosphate isomerase provides a direct link to the observed deglutathionylation of cysteine 217 in triosephosphate isomerase, thereby reversing the inhibitory effect of S-glutathionylation of triosephosphate isomerase.

The Structure of the Thioredoxin−Triosephosphate Isomerase Complex Provides Insights into the Reversible Glutathione-Mediate Regulation of Triosephosphate Isomerase

Protein−protein interactions are crucial for many biological functions. The redox interactome encompasses numerous weak transient interactions in which thioredoxin plays a central role. Proteomic studies have shown that thioredoxin binds to numerous proteins belonging to various cellular processes, including energy metabolism. Thioredoxin has cross talk with other redox mechanisms involving glutathionylation and has functional overlap with glutaredoxin in deglutathionylation reactions. In this study, we have explored the structural and biochemical interactions of thioredoxin with the glycolytic enzyme, triosephosphate isomerase. Nuclear magnetic resonance chemical shift mapping methods and molecular dynamics-based docking have been applied in deriving a structural model of the thioredoxin−triosephosphate isomerase complex. The spatial proximity of active site cysteine residues of thioredoxin to reactive thiol groups on triosephosphate isomerase provides a direct link to the observed deglutathionylation of cysteine 217 in triosephosphate isomerase, thereby reversing the inhibitory effect of S-glutathionylation of triosephosphate isomerase.

Insights into the Substrate Specificity of a Thioesterase Rv0098 of Mycobacterium Tuberculosis through X-ray Crystallographic and Molecular Dynamics Studies

The crystal structure of Rv0098, a long-chain fatty acyl-CoA thioesterase from Mycobacterium tuberculosis with bound dodecanoic acid at the active site provided insights into the mode of substrate binding but did not reveal the structural basis of substrate specificities of varying chain length. Molecular dynamics studies demonstrated that certain residues of the substrate binding tunnel are flexible and thus modulate the length of the tunnel. The flexibility of the loop at the base of the tunnel was also found to be important for determining the length of the tunnel for accommodating appropriate substrates. A combination of crystallographic and molecular dynamics studies thus explained the structural basis of accommodating long chain substrates by Rv0098 of M. tuberculosis.

Evaluation of the role of disordered organic fluorine in crystal packing: insights from halogen substituted benzanilides

The determination of the crystal and molecular structures of a large number of compounds containing the C(sp(2))-F bond has been investigated in detail in halogenated benzanilides and also in liquids, namely the fluorinated amines. It has been observed that when the fluorine atom is present in the ortho or meta position with respect to the amide functionality in benzanilides or the amino group in fluorinated amines which are liquids at room temperature, the fluorine atom exhibits positional disorder. This is associated with changes in patterns of intermolecular interactions which affect crystal packing. Furthermore, the presence of a fluorine atom on the benzanilide framework, in the presence of a heavier halogen (chloro, bromo and iodo), meta or ortho to the amide group does not eliminate the disorder associated with these molecules. In this article, we highlight the salient features present in halogenated compounds exhibiting disorder in the position of organic fluorine with concomitant changes in crystal packing. This feature is also compared with related compounds exhibiting similarity in electronic features, namely positional disorder.

Mechanistic insights into type III restriction enzymes

Type III restriction-modification (R-M) enzymes need to interact with two separate unmethylated DNA sequences in indirectly repeated, head-to-head orientations for efficient cleavage to occur at a defined location next to only one of the two sites. However, cleavage of sites that are not in head-to-head orientation have been observed to occur under certain reaction conditions in vitro. ATP hydrolysis is required for the long-distance communication between the sites prior to cleavage. Type III R-M enzymes comprise two subunits, Res and Mod that form a homodimeric Mod(2) and a heterotetrameric Res(2)Mod(2) complex. The Mod subunit in M-2 or R2M2 complex recognizes and methylates DNA while the Res subunit in R2M2 complex is responsible for ATP hydrolysis, DNA translocation and cleavage. A vast majority of biochemical studies on Type III R-M enzymes have been undertaken using two closely related enzymes, EcoP1I and EcoP15I. Divergent opinions about how the long-distance interaction between the recognition sites exist and at least three mechanistic models based on 1D- diffusion and/or 3D-DNA looping have been proposed.

Determinant: Old algorithms, new insights (Extended Abstract)

In this paper we approach the problem of computing the characteristic polynomial of a matrix from the combinatorial viewpoint. We present several combinatorial characterizations of the coefficients of the characteristic polynomial, in terms of walks and closed walks of different kinds in the underlying graph. We develop algorithms based on these characterizations, and show that they tally with well-known algorithms arrived at independently from considerations in linear algebra.

New Insights into Optimal Discrete Rate Adaptation for Average Power Constrained Single and Multi-Node Systems

The throughput-optimal discrete-rate adaptation policy, when nodes are subject to constraints on the average power and bit error rate, is governed by a power control parameter, for which a closed-form characterization has remained an open problem. The parameter is essential in determining the rate adaptation thresholds and the transmit rate and power at any time, and ensuring adherence to the power constraint. We derive novel insightful bounds and approximations that characterize the power control parameter and the throughput in closed-form. The results are comprehensive as they apply to the general class of Nakagami-m (m >= 1) fading channels, which includes Rayleigh fading, uncoded and coded modulation, and single and multi-node systems with selection. The results are appealing as they are provably tight in the asymptotic large average power regime, and are designed and verified to be accurate even for smaller average powers.

Insights into differential modulation of receptor function by hinge region using novel agonistic lutropin receptor and inverse agonistic thyrotropin receptor antibodies

We report two antibodies, scFv 13B1 and MAb PD1.37, against the hinge regions of LHR and TSHR, respectively, which have similar epitopes but different effects on receptor function. While neither of them affected hormone binding, with marginal effects on hormone response, scFv 13B1 stimulated LHR in a dose-dependent manner, whereas MAb PD1.37 acted as an inverse agonist of TSHR. Moreover, PD1.37 could decrease the basal activity of hinge region CAMs, but had varied effects on those present in ECLs, whereas 13B1 was refractory to any CAMs in LHR. Using truncation mutants and peptide phage display, we compared the differential roles of the hinge region cysteine box-2/3 as well as the exoloops in the activation of these two homologus receptors. (C) 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Structure and Mechanistic Insights into Novel Iron-mediated Moonlighting Functions of Human J-protein Cochaperone, Dph4

J-proteins are obligate cochaperones of Hsp70s and stimulate their ATPase activity via the J-domain. Although the functions of J-proteins have been well understood in the context of Hsp70s, their additional co-evolved ``physiological functions'' are still elusive. We report here the solution structure and mechanism of novel iron-mediated functional roles of human Dph4, a type III J-protein playing a vital role in diphthamide biosynthesis and normal development. The NMR structure of Dph4 reveals two domains: a conserved J-domain and a CSL-domain connected via a flexible linker-helix. The linker-helix modulates the conformational flexibility between the two domains, regulating thereby the protein function. Dph4 exhibits a unique ability to bind iron in tetrahedral coordination geometry through cysteines of its CSL-domain. The oxidized Fe-Dph4 shows characteristic UV-visible and electron paramagnetic resonance spectral properties similar to rubredoxins. Iron-bound Dph4 (Fe-Dph4) also undergoes oligomerization, thus potentially functioning as a transient ``iron storage protein,'' thereby regulating the intracellular iron homeostasis. Remarkably, Fe-Dph4 exhibits vital redox and electron carrier activity, which is critical for important metabolic reactions, including diphthamide biosynthesis. Further, we observed that Fe-Dph4 is conformationally better poised to perform Hsp70-dependent functions, thus underlining the significance of iron binding in Dph4. Yeast Jjj3, a functional ortholog of human Dph4 also shows a similar iron-binding property, indicating the conserved nature of iron sequestration across species. Taken together, our findings provide invaluable evidence in favor of additional co-evolved specialized functions of J-proteins, previously not well appreciated.

Insights into the Fold Organization of TIM Barrel from Interaction Energy Based Structure Networks

There are many well-known examples of proteins with low sequence similarity, adopting the same structural fold. This aspect of sequence-structure relationship has been extensively studied both experimentally and theoretically, however with limited success. Most of the studies consider remote homology or ``sequence conservation'' as the basis for their understanding. Recently ``interaction energy'' based network formalism (Protein Energy Networks (PENs)) was developed to understand the determinants of protein structures. In this paper we have used these PENs to investigate the common non-covalent interactions and their collective features which stabilize the TIM barrel fold. We have also developed a method of aligning PENs in order to understand the spatial conservation of interactions in the fold. We have identified key common interactions responsible for the conservation of the TIM fold, despite high sequence dissimilarity. For instance, the central beta barrel of the TIM fold is stabilized by long-range high energy electrostatic interactions and low-energy contiguous vdW interactions in certain families. The other interfaces like the helix-sheet or the helix-helix seem to be devoid of any high energy conserved interactions. Conserved interactions in the loop regions around the catalytic site of the TIM fold have also been identified, pointing out their significance in both structural and functional evolution. Based on these investigations, we have developed a novel network based phylogenetic analysis for remote homologues, which can perform better than sequence based phylogeny. Such an analysis is more meaningful from both structural and functional evolutionary perspective. We believe that the information obtained through the ``interaction conservation'' viewpoint and the subsequently developed method of structure network alignment, can shed new light in the fields of fold organization and de novo computational protein design.

Insights to urban dynamics through landscape spatial pattern analysis

Urbanisation is a dynamic complex phenomenon involving large scale changes in the land uses at local levels. Analyses of changes in land uses in urban environments provide a historical perspective of land use and give an opportunity to assess the spatial patterns, correlation, trends, rate and impacts of the change, which would help in better regional planning and good governance of the region. Main objective of this research is to quantify the urban dynamics using temporal remote sensing data with the help of well-established landscape metrics. Bangalore being one of the rapidly urbanising landscapes in India has been chosen for this investigation. Complex process of urban sprawl was modelled using spatio temporal analysis. Land use analyses show 584% growth in built-up area during the last four decades with the decline of vegetation by 66% and water bodies by 74%. Analyses of the temporal data reveals an increase in urban built up area of 342.83% (during 1973-1992), 129.56% (during 1992-1999), 106.7% (1999-2002), 114.51% (2002-2006) and 126.19% from 2006 to 2010. The Study area was divided into four zones and each zone is further divided into 17 concentric circles of 1 km incrementing radius to understand the patterns and extent of the urbanisation at local levels. The urban density gradient illustrates radial pattern of urbanisation for the period 1973-2010. Bangalore grew radially from 1973 to 2010 indicating that the urbanisation is intensifying from the central core and has reached the periphery of the Greater Bangalore. Shannon's entropy, alpha and beta population densities were computed to understand the level of urbanisation at local levels. Shannon's entropy values of recent time confirms dispersed haphazard urban growth in the city, particularly in the outskirts of the city. This also illustrates the extent of influence of drivers of urbanisation in various directions. Landscape metrics provided in depth knowledge about the sprawl. Principal component analysis helped in prioritizing the metrics for detailed analyses. The results clearly indicates that whole landscape is aggregating to a large patch in 2010 as compared to earlier years which was dominated by several small patches. The large scale conversion of small patches to large single patch can be seen from 2006 to 2010. In the year 2010 patches are maximally aggregated indicating that the city is becoming more compact and more urbanised in recent years. Bangalore was the most sought after destination for its climatic condition and the availability of various facilities (land availability, economy, political factors) compared to other cities. The growth into a single urban patch can be attributed to rapid urbanisation coupled with the industrialisation. Monitoring of growth through landscape metrics helps to maintain and manage the natural resources. (C) 2012 Elsevier B.V. All rights reserved.

High field carrier transport in graphene: Insights from fast current transient

In this work, we observe gate tunable negative differential conductance (NDC) and current saturation in single layer and bilayer graphene transistor at high source-drain field, which arise due to the interplay among (1) self-heating, (2) hot carrier injection, and (3) drain induced minority carrier injection. The magnitude of the NDC is found to be reduced for a bilayer, in agreement with its weaker carrier-optical phonon coupling and less efficient hot carrier injection. The contributions of different mechanisms to the observed results are decoupled through fast transient measurements with nanosecond resolution. The findings provide insights into high field transport in graphene. (C) 2012 American Institute of Physics. http://dx.doi.org/10.1063/1.4754103]

The antibodies against the computationally designed mimic of the glycoprotein hormone receptor transmembrane domain provide insights into receptor activation and suppress the constitutively activated receptor Mutants

The exoloops of glycoprotein hormone receptors (GpHRs) transduce the signal generated by the ligand-ectodomain interactions to the transmembrane helices either through direct hormonal contact and/or by modulating the interdomain interactions between the hinge region (HinR) and the transmembrane domain (TMD). The ligand-induced conformational alterations in the HinRs and the interhelical loops of luteinizing hormone receptor/follicle stimulating hormone receptor/thyroid stimulating hormone receptor were mapped using exoloop-specific antibodies generated against a mini-TMD protein designed to mimic the native exoloop conformations that were created by joining the thyroid stimulating hormone receptor exoloops constrained through helical tethers and library-derived linkers. The antibody against the mini-TMD specifically recognized all three GpHRs and inhibited the basal and hormone-stimulated cAMP production without affecting hormone binding. Interestingly, binding of the antibody to all three receptors was abolished by prior incubation of the receptors with the respective hormones, suggesting that the exoloops are buried in the hormone-receptor complexes. The antibody also suppressed the high basal activities of gain-of-function mutations in the HinRs, exoloops, and TMDs such as those involved in precocious puberty and thyroid toxic adenomas. Using the antibody and point/deletion/chimeric receptor mutants, we demonstrate that changes in the HinR-exoloop interactions play an important role in receptor activation. Computational analysis suggests that the mini-TMD antibodies act by conformationally locking the transmembrane helices by means of restraining the exoloops and the juxta-membrane regions. Using GpHRs as a model, we describe a novel computational approach of generating soluble TMD mimics that can be used to explain the role of exoloops during receptor activation and their interplay with TMDs.