Intra and Inter-Molecular Communications Through Protein Structure Network

Communication within and across proteins is crucial for the biological functioning of proteins. Experiments such as mutational studies on proteins provide important information on the amino acids, which are crucial for their function. However, the protein structures are complex and it is unlikely that the entire responsibility of the function rests on only a few amino acids. A large fraction of the protein is expected to participate in its function at some level or other. Thus, it is relevant to consider the protein structures as a completely connected network and then deduce the properties, which are related to the global network features. In this direction, our laboratory has been engaged in representing the protein structure as a network of non-covalent connections and we have investigated a variety of problems in structural biology, such as the identification of functional and folding clusters, determinants of quaternary association and characterization of the network properties of protein structures. We have also addressed a few important issues related to protein dynamics, such as the process of oligomerization in multimers, mechanism on protein folding, and ligand induced communications (allosteric effect). In this review we highlight some of the investigations which we have carried out in the recent past. A review on protein structure graphs was presented earlier, in which the focus was on the graphs and graph spectral properties and their implementation in the study of protein structure graphs/networks (PSN). In this article, we briefly summarize the relevant parts of the methodology and the focus is on the advancement brought out in the understanding of protein structure-function relationships through structure networks. The investigations of structural/biological problems are divided into two parts, in which the first part deals with the analysis of PSNs based on static structures obtained from x-ray crystallography. The second part highlights the changes in the network, associated with biological functions, which are deduced from the network analysis on the structures obtained from molecular dynamics simulations.

Crystallization behaviour of Metglas 2826 MB (Fe40Ni38Mo4B18)

Differential scanning calorimetry, x-ray diffraction and transmission electron microscopy have been employed to determine the thermal stability and identify the crystalline phases on devitrification of Metglas 2826 MB. The glass crystallizes intoγ-FeNiMo and fcc (FeNi)23B6 with activation energies of 270 and 375 kJ mol−1 respectively. The reactions are primary and polymorphic in nature. The influence of Mo towards crystallization of Fe40Ni40B20 has been to enhance the formation of the fcc (FeNi)23B6 phase in preference to orthorhombic (FeNi)3B phase and to raise the thermal stability of the amorphous state.

Influence of seeding on crystallization behaviour of BaNaB9O15 glasses

Transparent BaNaB9O15 (BNBO) glasses were fabricated via the conventional melt-quenching technique. X-ray powder diffraction (XRD) followed by differential scanning calorimetric (DSC) studies confirmed the amorphous and glassy nature of the as-quenched samples, respectively. The effect of seeding on the crystallization of BNBO glasses was studied by non-isothermal DSC method and was modeled using the Johnson-Mehl-Avrami and Ozawa equations. The activation energy for seeded glasses decreased with the increase in fraction of crystallization. The values for the onset of crystallization and Avrami exponent were found to be lower for seeded samples which were associated with the heterogeneous nucleation and epitaxial processes.

Crystallization studies on bulk SixTe100-x glasses

Bulk SixTe100-x (10 less-than-or-equals, slant x less-than-or-equals, slant 28) glasses have been prepared by the melt quenching technique. The crystallization of these glasses has been studied by using differential scanning calorimetry. The composition dependence of the glass transition temperature Tg, the recrystallization temperature Tcr and the melting temperature Tm show an unusual behaviour at the composition x congruent with 20. The glasses with 10 less-than-or-equals, slant x less-than-or-equals, slant 20 undergo double glass transition and double stage crystallization phenomena. On the other hand, glasses with 22 less-than-or-equals, slant x less-than-or-equals, slant 28 exhibit eutectic crystallization. The phases at different stages of crystallization have been identified by using X-ray diffraction techniques. The unusual behaviour at x congruent with 20 can be explained on the basis of the changes in the network topologies of IV-VI chalcogenide glasses.

Large cryptic internal sequence repeats in protein structures from Homo sapiens

Amino acid sequences are known to constantly mutate and diverge unless there is a limiting condition that makes such a change deleterious. However, closer examination of the sequence and structure reveals that a few large, cryptic repeats are nevertheless sequentially conserved. This leads to the question of why only certain repeats are conserved at the sequence level. It would be interesting to find out if these sequences maintain their conservation at the three-dimensional structure level. They can play an active role in protein and nucleotide stability, thus not only ensring proper functioning but also potentiating malfunction and disease. Therefore, insights into any aspect of the repeats - be it structure, function or evolution - would prove to be of some importance. This study aims to address the relationship between protein sequence and its three-dimensional structure, by examining if large cryptic sequence repeats have the same structure.

Allostery in tRNA Synthetases Elucidated from MD Simulations and Protein Structure Networks

tRNA synthetases (aaRS) are enzymes crucial in the translation of genetic code. The enzyme accylates the acceptor stem of tRNA by the congnate amino acid bound at the active site, when the anti-codon is recognized by the anti-codon site of aaRS. In a typical aaRS, the distance between the anti-codon region and the amino accylation site is approximately 70 Å. We have investigated this allosteric phenomenon at molecular level by MD simulations followed by the analysis of protein structure networks (PSN) of non-covalent interactions. Specifically, we have generated conformational ensembles by performing MD simulations on different liganded states of methionyl tRNA synthetase (MetRS) from Escherichia coli and tryptophenyl tRNA synthetase (TrpRS) from Human. The correlated residues during the MD simulations are identified by cross correlation maps. We have identified the amino acids connecting the correlated residues by the shortest path between the two selected members of the PSN. The frequencies of paths have been evaluated from the MD snapshots[1]. The conformational populations in different liganded states of the protein have been beautifully captured in terms of network parameters such as hubs, cliques and communities[2]. These parameters have been associated with the rigidity and plasticity of the protein conformations and can be associated with free energy landscape. A comparison of allosteric communication in MetRS and TrpRS [3] elucidated in this study highlights diverse means adopted by different enzymes to perform a similar function. The computational method described for these two enzymes can be applied to the investigation of allostery in other systems.

Structural Polymorphism of the Nucleosomal DNA and Implications for Protein Binding

A nucleosome forms a basic unit of the chromosome structure. A biologically relevant question is how much of the nucleosomal conformational space is accessible to protein-free DNA, and what proportion of the nucleosomal conformations are induced by bound histones. To investigate this, we have analysed high resolution xray crystal structure datasets of DNA in protein-free as well as protein-bound forms, and compared the dinucleotide step parameters for the two datasets with those for high resolution nucleosome structures. Our analysis shows that most of the dinucleotide step parameter values for the nucleosome structures lie within the range accessible to protein-free DNA, indirectly indicating that the histone core plays more of a stabilizing role. The nucleosome structures are observed to assume smooth and nearly planar curvature, implying that ‘normal’ B-DNA like parameters can give rise to a curved geometry at the gross structural level. Different nucleosome

Crystallization kinetics of poly(l-lactic acid)

The morphology and crystal growth of poly(l-lactic acid), PLLA have been studied from the melt as a function of undercooling and molecular weight using hot stage microscopy. Attention has been given to the application of growth rate equation on the growth rate data of PLLA and thus various nucleation parameters have been calculated. The criteria of Regime I and Regime II types of crystallization has been applied for the evaluation of substrate lengths.

Photo-induced double-strand DNA and site-specific protein cleavage activity of L-histidine (mu-oxo)diiron(III) complexes of heterocyclic bases

Three oxo-bridged diiron(III) complexes of L-histidine and heterocyclic bases [Fe-2(mu-O)(L-his)(2)(B)(2)](ClO4)(2) (1-3), where B is 2,2'-bipyridine (bpy),1,10-phenanthroline (phen), dipyrido[3,2-d:2',3'-f]quinoxaline (dpq), were prepared and characterized. The bpy complex 1 was structurally characterized by X-ray crystallography. The molecular structure showed a {Fe-2(mu-O)} core in which iron(III) in a FeN4O2 coordination is bound to tridentate monoanionic L-histidine and bidentate bpy ligands. The Fe center dot center dot center dot Fe distance is similar to 3.5 angstrom. The Fe-O-Fe unit is essentially linear, giving a bond angle of similar to 172 degrees. The complexes showed irreversible cyclic voltammetric cathodic response near -0.1 V vs. SCE in H2O-0.1 M KCl. The binuclear units displayed antiferromagnetic interaction between two high-spin (S = 5/2) iron(III) centers giving a -J value of -110 cm(-1). The complexes showed good DNA binding propensity giving a binding constant value of similar to 10(5) M-1. Isothermal titration calorimetric data indicated single binding mode to the DNA. The binding was found to be driven by negative free energy change and enthalpy. The dpq complex 3 showed oxidative double-strand DNA cleavage on exposure to UV-A and visible light. The phen complex 2 displayed single-strand photocleavage of DNA. The DNA double-strand breaks were rationalized from theoretical molecular docking calculations. Mechanistic investigations showed formation of hydroxyl radicals as the reactive species through photodecarboxylation of the L-histidine ligand. The complexes exhibited good binding propensity to bovine serum albumin (BSA) protein in Tris-HCl/NaCl buffer medium. The dpq complex 3 showed UV-A light-induced site-specific oxidative BSA cleavage forming fragments of similar to 45 kDa and similar to 20 kDa molecular weights via SOH pathway.

Small local variations in B-form DNA lead to a large variety of global geometries which can accommodate most DNA-binding protein motifs

An important question of biological relevance is the polymorphism of the double-helical DNA structure in its free form, and the changes that it undergoes upon protein-binding. We have analysed a database of free DNA crystal structures to assess the inherent variability of the free DNA structure and have compared it with a database of protein-bound DNA crystal structures to ascertain the protein-induced variations.

Estrogen modulation of retinol-binding protein in immature chicks: Comparison with riboflavin carrier protein

The kinetics of estrogen (E) modulation of retinol-binding protein (RBP) production in the liver of immature chicks were compared with those governing de novo induction of riboflavin carrier protein (RCP) in the same tissue. A single dose of E markedly enhanced the plasma levels of RBP without any detectable lag period to reach peak value by 24 h and this was followed by a decline to attain the baseline by 4 days. There was no amplification of the response during secondary stimulation unlike the case with RCP induction. With multiple E administration, the 4-fold increased plasma RBP concentrations were sustained at a steady state during both primary and secondary stimulations, whereas concomitant RCP concentration progressively increased with each hormone administration and this response was further amplified during secondary stimulation. Unlike RCP induction, enhanced RBP accumulation was not strictly E dose dependent although a minimal threshold level of the steroid was required to elicit measurable response. Progesterone (P) could neither modulate nor substitute for E in enhancing plasma levels of either of the 2 proteins while the anti-estrogens, en- and zuclomifene citrate severely suppressed the production of both the proteins. RCP induction was completely inhibited by both α-amanitin and cycloheximide for prolonged periods while E-stimulated RBP production was affected only partially by α-amanitin. Likewise, cycloheximide inhibition of RBP accumulation followed a pattern similar to that of hepatic general protein synthesis.