Narayanaswamy Srinivasan

PIC: Protein Interactions Calculator

Interactions within a protein structure and interactions between proteins in an assembly are essential considerations in understanding molecular basis of stability and functions of proteins and their complexes. There are several weak and strong interactions that render stability to a protein structure or an assembly. Protein Interactions Calculator (PIC) is a server which, given the coordinate set of 3D structure of a protein or an assembly, computes various interactions such as disulphide bonds, interactions between hydrophobic residues, ionic interactions, hydrogen bonds, aromatic–aromatic interactions, aromatic–sulphur interactions and cation–π interactions within a protein or between proteins in a complex. Interactions are calculated on the basis of standard, published criteria. The identified interactions between residues can be visualized using a RasMol and Jmol interface. The advantage with PIC server is the easy availability of inter-residue interaction calculations in a single site. It also determines the accessible surface area and residue-depth, which is the distance of a residue from the surface of the protein. User can also recognize specific kind of interactions, such as apolar–apolar residue interactions or ionic interactions, that are formed between buried or exposed residues or near the surface or deep inside.

Knowledge-Based Protein Modeling

Knowledge, both from the three-dimensional structures of homologous proteins and from the general analysis of protein structure, is of value in modeling a protein of known sequence but unknown structure. While many models are still constructed at least in part by manual methods on graphics devices, automated procedures have come into greater use. These procedures include those that assemble fragments of structure from other known structures and those that derive coordinates for the model from the satisfaction of restraints placed on atomic positions.

A genomic perspective of protein kinases in Plasmodium falciparum

Protein kinases are central to regulation of cellular signaling in the eukaryotes. Well‐conserved and lineage‐specific protein kinases have previously been identified from various completely sequenced genomes of eukaryotes. The current work describes a genome‐wide analysis for protein kinases encoded in the Plasmodium falciparum genome. Using a few different profile matching methods, we have identified 99 protein kinases or related proteins in the parasite genome. We have classified these kinases into subfamilies and analyzed them in the context of noncatalytic domains that occur in these catalytic kinase domain–containing proteins. Compared to most eukaryotic protein kinases, these sequences vary significantly in terms of their lengths, inserts in catalytic domains, and co‐occurring domains. Catalytic and noncatalytic domains contain long stretches of repeats of positively charged and other polar amino acids. Various components of the cell cycle, including 4 cyclin‐dependent kinase (CDK) homologues, 2 cyclins, 1 CDK regulatory subunit, and 1 kinase‐associated phosphatase, are identified. Identification of putative mitogen‐activated protein (MAP) Kinase and MAP Kinase Kinase of P. falciparum suggests a new paradigm in the highly conserved signaling pathway of eukaryotes. The calcium‐dependent kinase family, well represented in P. falciparum, shows varying domain combinations with EF‐hands and pleckstrin homology domains. The analysis reveals a new subfamily of protein kinases having limited sequence similarity with previously known subfamilies. A new transmembrane kinase with 6 membrane‐spanning regions is identified. Putative apicoplast targeting sequences have been detected in some of these protein kinases, suggesting their export to the apicoplast. Proteins 2005.

Interaction preferences across protein-protein interfaces of obligatory and non-obligatory components are different

BackgroundA polypeptide chain of a protein-protein complex is said to be obligatory if it is bound to another chain throughout its functional lifetime. Such a chain might not adopt the native fold in the unbound form. A non-obligatory polypeptide chain associates with another chain and dissociates upon molecular stimulus. Although conformational changes at the interaction interface are expected, the overall 3-D structure of the non-obligatory chain is unaltered. The present study focuses on protein-protein complexes to understand further the differences between obligatory and non-obligatory interfaces.ResultsA non-obligatory chain in a complex of known 3-D structure is recognized by its stable existence with same fold in the bound and unbound forms. On the contrary, an obligatory chain is detected by its existence only in the bound form with no evidence for the native-like fold of the chain in the unbound form. Various interfacial properties of a large number of complexes of known 3-D structures thus classified are comparatively analyzed with an aim to identify structural descriptors that distinguish these two types of interfaces. We report that the interaction patterns across the interfaces of obligatory and non-obligatory components are different and contacts made by obligatory chains are predominantly non-polar. The obligatory chains have a higher number of contacts per interface (20 ± 14 contacts per interface) than non-obligatory chains (13 ± 6 contacts per interface). The involvement of main chain atoms is higher in the case of obligatory chains (16.9 %) compared to non-obligatory chains (11.2 %). The β-sheet formation across the subunits is observed only among obligatory protein chains in the dataset. Apart from these, other features like residue preferences and interface area produce marginal differences and they may be considered collectively while distinguishing the two types of interfaces.ConclusionThese results can be useful in distinguishing the two types of interfaces observed in structures determined in large-scale in the structural genomics initiatives, especially for those multi-component protein assemblies for which the biochemical characterization is incomplete.

Comparative analyses of pentraxins: implications for protomer assembly and ligand binding.

BACKGROUND Pentraxins are a family of plasma proteins characterized by their pentameric assembly and calcium-dependent ligand binding. The recent determination of the crystal structure for a member of this family, human serum amyloid P component (SAP), provides a basis for the comparative analysis of the pentraxin family. RESULTS We have compared the sequences, tertiary structures and quaternary arrangements of SAP with human C-reactive protein (CRP), Syrian hamster SAP (HSAP) and Limulus polyphemus CRP (LIM). These proteins can adopt a beta-jelly roll topology and a hydrophobic core similar to that seen in SAP. Only minor differences are observed in the positions of residues involved in coordinating calcium ions. CONCLUSIONS Calcium-mediated ligand binding by CRP, HSAP and LIM is similar to that defined by the crystal structure of SAP, but sequence differences in the hydrophobic pocket explain the differential ligand specificities exhibited by the homologous proteins. Differences elsewhere, including insertions and deletions, account for the different (hexameric) quaternary structure of LIM.

Identification of specific DNA binding residues in the TCP family of transcription factors in Arabidopsis.

The TCP family is a group of plant-specific transcription factors whose DNA binding properties have not been studied in detail. Here, we examine TCP4 by both biochemical and structural analyses to describe the DNA binding mechanisms of this family of proteins and predict a fold that the domain might adopt. The TCP transcription factors control multiple developmental traits in diverse plant species. Members of this family share an ∼60-residue-long TCP domain that binds to DNA. The TCP domain is predicted to form a basic helix-loop-helix (bHLH) structure but shares little sequence similarity with canonical bHLH domain. This classifies the TCP domain as a novel class of DNA binding domain specific to the plant kingdom. Little is known about how the TCP domain interacts with its target DNA. We report biochemical characterization and DNA binding properties of a TCP member in Arabidopsis thaliana, TCP4. We have shown that the 58-residue domain of TCP4 is essential and sufficient for binding to DNA and possesses DNA binding parameters comparable to canonical bHLH proteins. Using a yeast-based random mutagenesis screen and site-directed mutants, we identified the residues important for DNA binding and dimer formation. Mutants defective in binding and dimerization failed to rescue the phenotype of an Arabidopsis line lacking the endogenous TCP4 activity. By combining structure prediction, functional characterization of the mutants, and molecular modeling, we suggest a possible DNA binding mechanism for this class of transcription factors.

Hepatitis C virus internal ribosome entry site-mediated translation is stimulated by specific interaction of independent regions of human La autoantigen

The human La autoantigen has been shown to interact with the internal ribosome entry site (IRES) of hepatitis C virus (HCV) in vitro. Using a yeast three-hybrid system, we demonstrated that, in addition to full-length La protein, both N- and C-terminal halves were able to interact with HCV IRES in vivo. The exogenous addition of purified full-length and truncated La proteins in rabbit reticulocyte lysate showed dose-dependent stimulation of HCV IRES-mediated translation. However, an additive effect was achieved adding the terminal halves together in the reaction, suggesting that both might play critical roles in achieving full stimulatory activity of the full-length La protein. Using computational analysis, three-dimensional structures of the RNA recognition motifs (RRM) of the La protein were independently modeled. Of the three putative RRMs, RRM2 was predicted to have a good binding pocket for the interaction with the HCV IRES around the GCAC motif near the initiator AUG and RRM3 binds perhaps in a different location. This observation was further investigated by the filter-binding and toe-printing assays. The results presented here strongly suggest that both the N- and C-terminal halves can interact independently with the HCV IRES and are involved in stimulating internal initiation of translation.

iPBA: a tool for protein structure comparison using sequence alignment strategies

With the immense growth in the number of available protein structures, fast and accurate structure comparison has been essential. We propose an efficient method for structure comparison, based on a structural alphabet. Protein Blocks (PBs) is a widely used structural alphabet with 16 pentapeptide conformations that can fairly approximate a complete protein chain. Thus a 3D structure can be translated into a 1D sequence of PBs. With a simple Needleman–Wunsch approach and a raw PB substitution matrix, PB-based structural alignments were better than many popular methods. iPBA web server presents an improved alignment approach using (i) specialized PB Substitution Matrices (SM) and (ii) anchor-based alignment methodology. With these developments, the quality of ∼88% of alignments was improved. iPBA alignments were also better than DALI, MUSTANG and GANGSTA+ in >80% of the cases. The webserver is designed to for both pairwise comparisons and database searches. Outputs are given as sequence alignment and superposed 3D structures displayed using PyMol and Jmol. A local alignment option for detecting subs-structural similarity is also embedded. As a fast and efficient ‘sequence-based’ structure comparison tool, we believe that it will be quite useful to the scientific community. iPBA can be accessed at http://www.dsimb.inserm.fr/dsimb_tools/ipba/.

CAMPASS: a database of structurally aligned protein superfamilies

We thank Andrej Sali, John Overington and Mark Johnson for use of the programs COMPARER, JOY and MALIGN, respectively. We thank the anonymous referees for their useful comments. For financial support, RS thanks the Imperial Cancer Research Fund; DFB and HAN Oxford Molecular Ltd; KM HFSP; NS The Wellcome Trust; and RES the MRC.

KinG: a database of protein kinases in genomes

The KinG database is a comprehensive collection of serine/threonine/tyrosine-specific kinases and their homologues identified in various completed genomes using sequence and profile search methods. The database hosted at http://hodgkin. mbu.iisc.ernet.in/ approximately king provides the amino acid sequences, functional domain assignments and classification of gene products containing protein kinase domains. A search tool enabling the retrieval of protein kinases with specified subfamily and domain combinations is one of the key features of the resource. Identification of a kinase catalytic domain in the users query sequence is possible using another search tool. The occurrence and location of critical catalytic residues if the query has a catalytic kinase domain, recognition of non-kinase domains in the sequence and subfamily classification of the kinase in the query will help in deciphering the biological role of the kinase. This online compilation can also be used to compare the protein kinases of a given subfamily and domain combinations across various genomes. Another exclusive feature of the database is the collection of the Ser/Thr/Tyr protein kinases and similar sequences encoded in the genomes of archaea and bacteria.