Application of Computational Molecular Biophysics to Problems in Bacterial Chemotaxis

The combination of physics, biology, chemistry, and computer science constitutes the promising field of computational molecular biophysics. This field studies the molecular properties of DNA, protein lipids and biomolecules using computational methods. For this dissertation, I approached four problems involving the chemotaxis pathway, the set of proteins that function as the navigation system of bacteria and lower eukaryotes. In the first chapter, I used a special-purpose machine for molecular dynamics simulations, Anton, to simulate the signaling domain of the chemoreceptor in different signaling states for a total of 6 microseconds. Among other findings, this study provides enough evidence to propose a novel molecular mechanism for the kinase activation by the chemoreceptor and reconcile previously conflicting experimental data. In the second chapter, my molecular dynamics studies of the scaffold protein cheW reveals the existence and role of a conserved salt-bridge that stabilizes the relative position of the two binding sites in the chew surface: the chemoreceptor and the kinase. The results were further confirmed with NMR experiments performed with collaborators at the University of California in Santa Barbara, CA. In the third chapter, my colleagues and I investigate the quality of homology modeled structures with cheW protein as a benchmark. By subjecting the models to molecular dynamics and Monte Carlo simulations, we show that the homology models are snapshots of a larger ensemble of conformations very similar to the one generated by the experimental structures. In the fourth chapter, I use bioinformatics and basic mathematical modeling to predict the specific chemoreceptor(s) expressed in vivo and imaged with electron cryo tomography (ECT) by our collaborators at the California Institute of Technology. The study was essential to validate the argument that the hexagonal arrangement of transmembrane chemoreceptors is universal among bacteria, a major breakthrough in the field of chemotaxis. In summary, this thesis presents a collection of four works in the field of bacterial chemotaxis where either methods of physics or the quantitative approach of physicists were of fundamental importance for the success of the project

Evolutionary Genomics Suggests That CheV Is an Additional Adaptor for Accommodating Specific Chemoreceptors within the Chemotaxis Signaling Complex

Escherichia coli and Salmonella enterica are models for many experiments in molecular biology including chemotaxis, and most of the results obtained with one organism have been generalized to another. While most components of the chemotaxis pathway are strongly conserved between the two species, Salmonella genomes contain some chemoreceptors and an additional protein, CheV, that are not found in E. coli. The role of CheV was examined in distantly related species Bacillus subtilis and Helicobacter pylori, but its role in bacterial chemotaxis is still not well understood. We tested a hypothesis that in enterobacteria CheV functions as an additional adaptor linking the CheA kinase to certain types of chemoreceptors that cannot be effectively accommodated by the universal adaptor CheW. Phylogenetic profiling, genomic context and comparative protein sequence analyses suggested that CheV interacts with specific domains of CheA and chemoreceptors from an orthologous group exemplified by the Salmonella McpC protein. Structural consideration of the conservation patterns suggests that CheV and CheW share the same binding spot on the chemoreceptor structure, but have some affinity bias towards chemoreceptors from different orthologous groups. Finally, published experimental results and data newly obtained via comparative genomics support the idea that CheV functions as a “phosphate sink” possibly to off-set the over-stimulation of the kinase by certain types of chemoreceptors. Overall, our results strongly suggest that CheV is an additional adaptor for accommodating specific chemoreceptors within the chemotaxis signaling complex

Regular Architecture (RegArch): A standard expression language for describing protein architectures

Domain architecture – the arrangement of features in a protein – exhibits syntactic patterns similar to the grammar of a language. This feature enables pattern mining for protein function prediction, comparative genomics, and studies of molecular evolution and complexity. To facilitate such work, here we propose Regular Architecture (RegArch), an expression language to describe syntactic patterns in protein architectures. Like the well-known Regular Expressions for text, RegArchs codify positional and non-positional patterns of elements into nested JSON objects. We describe the standard and provide a reference implementation in JavaScript to parse RegArchs and match annotated proteins

Chemotaxis cluster 1 proteins form cytoplasmic arrays in Vibrio cholera and are stabilized by a double signaling domain receptor DosM

Nearly all motile bacterial cells use a highly sensitive and adaptable sensory system to detect changes in nutrient concentrations in the environment and guide their movements toward attractants and away from repellents. The best-studied bacterial chemoreceptor arrays are membrane-bound. Many motile bacteria contain one or more additional, sometimes purely cytoplasmic, chemoreceptor systems. Vibrio cholerae contains three chemotaxis clusters (I, II, and III). Here, using electron cryotomography, we explore V. cholerae’s cytoplasmic chemoreceptor array and establish that it is formed by proteins from cluster I. We further identify a chemoreceptor with an unusual domain architecture, DosM, which is essential for formation of the cytoplasmic arrays. DosM contains two signaling domains and spans the two-layered cytoplasmic arrays. Finally, we present evidence suggesting that this type of receptor is important for the structural stability of the cytoplasmic array

MiST 3.0: an updated microbial signal transduction database with an emphasis on chemosensory systems

Bacteria and archaea employ dedicated signal transduction systems that modulate gene expression, second-messenger turnover, quorum sensing, biofilm formation, motility, host-pathogen and beneficial interactions. The updated MiST database provides a comprehensive classification of microbial signal transduction systems. This update is a result of a substantial scaling to accommodate constantly growing microbial genomic data. More than 125 000 genomes, 516 million genes and almost 100 million unique protein sequences are currently stored in the database. For each bacterial and archaeal genome, MiST 3.0 provides a complete signal transduction profile, thus facilitating theoretical and experimental studies on signal transduction and gene regulation. Newsoftware infrastructure and distributed pipeline implemented in MiST 3.0 enable regular genome updates based on the NCBI RefSeq database. A novel MiST feature is the integration of unique profile HMMs to link complex chemosensory systems with corresponding chemoreceptors in bacterial and archaeal genomes