[Analysis of Multiple Protein Alignments Using 3D-Structural Information on the Orientation of Amino Acid Side-Chains].

Multiple alignment of amino acid sequences of homologous proteins is a key tool in state-of-the-art bioinformatics and evolutionary analysis. Differences in the spatial orientation of amino acid side-chains can predetermine significant functional diversity among members of one superfamily; however, this is usually not taken into account in any way when constructing alignments and during subsequent comparative analysis. First of all, this is due to the limitation of existing algorithms, which are guided by the biochemical similarity of the "alphabet" of amino acid substitutions and either do not use information about the 3D-structural organization of proteins at all, or are limited to comparing the backbone only (i.

Loop 422-437 in NanA from Streptococcus pneumoniae plays the role of an active site lid and is associated with allosteric regulation.

Neuraminidase A from Streptococcus pneumoniae (NanA) is considered a potentially key pathogenicity factor and a promising drug target to treat human infectious diseases. Computational and experimental efforts are increasingly being used to study its structure and function which yet remain poorly understood. In this work, we characterized structural dynamics of NanA's active site and gained novel mechanistic insights into its implications for a ligand binding.

Guide tree optimization with genetic algorithm to improve multiple protein 3D-structure alignment.

Motivation: With the increasing availability of 3D-data, the focus of comparative bioinformatic analysis is shifting from protein sequence alignments toward more content-rich 3D-alignments. This raises the need for new ways to improve the accuracy of 3D-superimposition.

Results: We proposed guide tree optimization with genetic algorithm (GA) as a universal tool to improve the alignment quality of multiple protein 3D-structures systematically.

Probing the role of the residues in the active site of the transaminase from Thermobaculum terrenum.

Creating biocatalysts for (R)-selective amination effectively is highly desirable in organic synthesis. Despite noticeable progress in the engineering of (R)-amine activity in pyridoxal-5'-phosphate-dependent transaminases of fold type IV, the specialization of the activity is still an intuitive task, as there is poor understanding of sequence-structure-function relationships. In this study, we analyzed this relationship in transaminase from Thermobaculum terrenum, distinguished by expanded substrate specificity and activity in reactions with L-amino acids and (R)-(+)-1-phenylethylamine using α-ketoglutarate and pyruvate as amino acceptors.

Bioinformatic Analysis of the Nicotinamide Binding Site in Poly(ADP-Ribose) Polymerase Family Proteins.

The PARP family consists of 17 members with diverse functions, including those related to cancer cells' viability. Several PARP inhibitors are of great interest as innovative anticancer drugs, but they have low selectivity towards distinct PARP family members and exert serious adverse effects. We describe a family-wide study of the nicotinamide (NA) binding site, an important functional region in the PARP structure, using comparative bioinformatic analysis and molecular modeling.

Bioinformatic analysis of subfamily-specific regions in 3D-structures of homologs to study functional diversity and conformational plasticity in protein superfamilies.

Local 3D-structural differences in homologous proteins contribute to functional diversity observed in a superfamily, but so far received little attention as bioinformatic analysis was usually carried out at the level of amino acid sequences. We have developed Zebra3D - the first-of-its-kind bioinformatic software for systematic analysis of 3D-alignments of protein families using machine learning. The new tool identifies subfamily-specific regions (SSRs) - patterns of local 3D-structure (i.

Mustguseal and Sister Web-Methods: A Practical Guide to Bioinformatic Analysis of Protein Superfamilies.

Bioinformatic analysis of functionally diverse superfamilies can help to study the structure-function relationship in proteins, but represents a methodological challenge. The Mustguseal web-server can build large structure-guided sequence alignments of thousands of homologs that cover all currently available sequence variants within a common structural fold. The input to the method is a PDB code of the query protein, which represents the protein superfamily of interest.

Catalytic and lectin domains in neuraminidase A from Streptococcus pneumoniae are capable of an intermolecular assembly: Implications for biofilm formation.

Neuraminidase A from Streptococcus pneumoniae (NanA) is a cell wall-bound modular enzyme containing one lectin and one catalytic domain. Unlike homologous NanB and NanC expressed by the same bacterium, the two domains within one NanA molecule do not form a stable interaction and are spatially separated by a 16-amino acid-long flexible linker. In this work, the ability of NanA to form intermolecular assemblies was characterized using the methods of molecular modeling and bioinformatic analysis based on crystallographic data and by bringing together previously published experimental data.

EasyAmber: A comprehensive toolbox to automate the molecular dynamics simulation of proteins.

Conformational plasticity of the functionally important regions and binding sites in protein/enzyme structures is one of the key factors affecting their function and interaction with substrates/ligands. Molecular dynamics (MD) can address the challenge of accounting for protein flexibility by predicting the time-dependent behavior of a molecular system. It has a potential of becoming a particularly important tool in protein engineering and drug discovery, but requires specialized training and skills, what impedes practical use by many investigators.

vsFilt: A Tool to Improve Virtual Screening by Structural Filtration of Docking Poses.

The ability of ligands to form crucial interactions with a protein target, characteristic for the substrate and/or inhibitors, could be considered a structural criterion for identifying potent binders among docked compounds. Structural filtration of predicted poses improves the performance of virtual screening and helps in recovering specifically bound ligands. Here, we present vsFilt-a highly automated and easy-to-use Web server for postdocking structural filtration.

Zebra2: advanced and easy-to-use web-server for bioinformatic analysis of subfamily-specific and conserved positions in diverse protein superfamilies.

Zebra2 is a highly automated web-tool to search for subfamily-specific and conserved positions (i.e. the determinants of functional diversity as well as the key catalytic and structural residues) in protein superfamilies.

Yosshi: a web-server for disulfide engineering by bioinformatic analysis of diverse protein families.

Disulfide bonds play a significant role in protein stability, function or regulation but are poorly conserved among evolutionarily related proteins. The Yosshi can help to understand the role of S-S bonds by comparing sequences and structures of homologs with diverse properties and different disulfide connectivity patterns within a common structural fold of a superfamily, and assist to select the most promising hot-spots to improve stability of proteins/enzymes or modulate their functions by introducing naturally occurring crosslinks. The bioinformatic analysis is supported by the integrated Mustguseal web-server to construct large structure-guided sequence alignments of functionally diverse protein families that can include thousands of proteins based on all available information in public databases.

CASBench: A Benchmarking Set of Proteins with Annotated Catalytic and Allosteric Sites in Their Structures.

In recent years, the phenomenon of allostery has witnessed growing attention driven by a fundamental interest in new ways to regulate the functional properties of proteins, as well as the prospects of using allosteric sites as targets to design novel drugs with lower toxicity due to a higher selectivity of binding and specificity of the mechanism of action. The currently available bioinformatic methods can sometimes correctly detect previously unknown ligand binding sites in protein structures. However, the development of universal and more efficient approaches requires a deeper understanding of the common and distinctive features of the structural organization of both functional (catalytic) and allosteric sites, the evolution of their amino acid sequences in respective protein families, and allosteric communication pathways.

parMATT: parallel multiple alignment of protein 3D-structures with translations and twists for distributed-memory systems.

Motivation: Accurate structural alignment of proteins is crucial at studying structure-function relationship in evolutionarily distant homologues. Various software tools were proposed to align multiple protein 3D-structures utilizing one CPU and thus are of limited productivity at large-scale analysis of protein families/superfamilies.

Results: The parMATT is a hybrid MPI/pthreads/OpenMP parallel re-implementation of the MATT algorithm to align multiple protein 3D-structures by allowing translations and twists.

Biochemical and structural insights into PLP fold type IV transaminase from Thermobaculum terrenum.

The high catalytic efficiency of enzymes under reaction conditions is one of the main goals in biocatalysis. Despite the dramatic progress in the development of more efficient biocatalysts by protein design, the search for natural enzymes with useful properties remains a promising strategy. The pyridoxal 5'-phosphate (PLP)-dependent transaminases represent a group of industrially important enzymes due to their ability to stereoselectively transfer amino groups between diverse substrates; however, the complex mechanism of substrate recognition and conversion makes the design of transaminases a challenging task.

Bioinformatic analysis of the fold type I PLP-dependent enzymes reveals determinants of reaction specificity in l-threonine aldolase from .

Understanding the role of specific amino acid residues in the molecular mechanism of a protein's function is one of the most challenging problems in modern biology. A systematic bioinformatic analysis of protein families and superfamilies can help in the study of structure-function relationships and in the design of improved variants of enzymes/proteins, but represents a methodological challenge. The pyridoxal-5'-phosphate (PLP)-dependent enzymes are catalytically diverse and include the aspartate aminotransferase superfamily which implements a common structural framework known as type fold I.

Human p38α mitogen-activated protein kinase in the Asp168-Phe169-Gly170-in (DFG-in) state can bind allosteric inhibitor Doramapimod.

Doramapimod (BIRB-796) is widely recognized as one of the most potent and selective type II inhibitors of human p38α mitogen-activated protein kinase (MAPK); however, the understanding of its binding mechanism remains incomplete. Previous studies indicated high affinity of the ligand to a so-called allosteric pocket revealed only in the 'out' state of the DFG motif (i.e.

Neuraminidase A from Streptococcus pneumoniae has a modular organization of catalytic and lectin domains separated by a flexible linker.

Neuraminidase A (NanA) of the pathogen Streptococcus pneumoniae cleaves receptors of the human respiratory epithelial surface during bacterial colonization. The full-size structure of NanA that contains one lectin and one catalytic domain within a single polypeptide chain remains unresolved. Both domains are crucial for the microorganism's virulence and considered as promising antimicrobial targets.

The visualCMAT: A web-server to select and interpret correlated mutations/co-evolving residues in protein families.

The visualCMAT web-server was designed to assist experimental research in the fields of protein/enzyme biochemistry, protein engineering, and drug discovery by providing an intuitive and easy-to-use interface to the analysis of correlated mutations/co-evolving residues. Sequence and structural information describing homologous proteins are used to predict correlated substitutions by the Mutual information-based CMAT approach, classify them into spatially close co-evolving pairs, which either form a direct physical contact or interact with the same ligand (e.g.

Mustguseal: a server for multiple structure-guided sequence alignment of protein families.

Motivation: Comparative analysis of homologous proteins in a functionally diverse superfamily is a valuable tool at studying structure-function relationship, but represents a methodological challenge.

Results: The Mustguseal web-server can automatically build large structure-guided sequence alignments of functionally diverse protein families that include thousands of proteins basing on all available information about their structures and sequences in public databases. Superimposition of protein structures is implemented to compare evolutionarily distant relatives, whereas alignment of sequences is used to compare close homologues.

Experimental and computational studies on the unusual substrate specificity of branched-chain amino acid aminotransferase from Thermoproteus uzoniensis.

PLP-Dependent fold-type IV branched-chain amino acid aminotransferases (BCATs) from archaea have so far been poorly characterized. A new BCAT from the hyperthermophilic archaeon Thermoproteus uzoniensis (TUZN1299) has been studied. TUZN1299 was found to be highly active toward branched-chain amino acids (BCAAs), positively charged amino acids, l-methionine, l-threonine, l-homoserine, l-glutamine, as well as toward 2-oxobutyrate and keto analogs of BCAAs, whereas l-glutamate and α-ketoglutarate were not converted in the overall reaction.

Parallel workflow manager for non-parallel bioinformatic applications to solve large-scale biological problems on a supercomputer.

Rapid expansion of online resources providing access to genomic, structural, and functional information associated with biological macromolecules opens an opportunity to gain a deeper understanding of the mechanisms of biological processes due to systematic analysis of large datasets. This, however, requires novel strategies to optimally utilize computer processing power. Some methods in bioinformatics and molecular modeling require extensive computational resources.

Study of Functional and Allosteric Sites in Protein Superfamilies.

The interaction of proteins (enzymes) with a variety of low-molecular-weight compounds, as well as protein-protein interactions, is the most important factor in the regulation of their functional properties. To date, research effort has routinely focused on studying ligand binding to the functional sites of proteins (active sites of enzymes), whereas the molecular mechanisms of allosteric regulation, as well as binding to other pockets and cavities in protein structures, remained poorly understood. Recent studies have shown that allostery may be an intrinsic property of virtually all proteins.

Robust enzyme design: bioinformatic tools for improved protein stability.

The ability of proteins and enzymes to maintain a functionally active conformation under adverse environmental conditions is an important feature of biocatalysts, vaccines, and biopharmaceutical proteins. From an evolutionary perspective, robust stability of proteins improves their biological fitness and allows for further optimization. Viewed from an industrial perspective, enzyme stability is crucial for the practical application of enzymes under the required reaction conditions.

Computational design of a pH stable enzyme: understanding molecular mechanism of penicillin acylase's adaptation to alkaline conditions.

Protein stability provides advantageous development of novel properties and can be crucial in affording tolerance to mutations that introduce functionally preferential phenotypes. Consequently, understanding the determining factors for protein stability is important for the study of structure-function relationship and design of novel protein functions. Thermal stability has been extensively studied in connection with practical application of biocatalysts.