How is structural divergence related to evolutionary information?

The analysis of evolutionary information in a protein family, such as conservation and covariation, is often linked to its structural information. Multiple sequence alignments of distant homologous sequences are used to measure evolutionary variables. Although high structural differences between proteins can be expected in such divergent alignments, most works linking evolutionary and structural information use a single structure ignoring the structural variability within protein families. The goal of this work is to elucidate the relevance of structural divergence when sequence-based measures are integrated with structural information. We found that inter-residue contacts and solvent accessibility undergo large variations in protein families. Our results show that high covariation scores tend to reveal residue contacts that are conserved in the family, instead of protein or conformer specific contacts. We also found that residue accessible surface area shows a high variability between structures of the same family. As a consequence, the mean relative solvent accessibility of multiple structures correlates better with the conservation pattern than the relative solvent accessibility of a single structure. We conclude that the use of comprehensive structural information allows a more accurate interpretation of the information computed from sequence alignments. Therefore, considering structural divergence would lead to a better understanding of protein function, dynamics, and evolution.Fil: Zea, Diego Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Monzón, Alexander. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Parisi, Gustavo Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Marino, Cristina Ester. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin

Probing structure, function and dynamics in bacterial primary and secondary transporter-associated binding proteins

Substrate binding proteins (SBPs) are ubiquitous in all life forms and have evolved to perform diverse physiological functions, such as in membrane transport, gene regulation, neurotransmission, and quorum sensing. It is quite astounding to observe such functional diversity among the SBPs even when they are restricted by their fold space. Therefore, the SBPs are an excellent set of proteins that can reveal how proteins evolution novel function in a structurally conserved/constrained fold. This study attempts to understand the phenomenon of affinity and specificity evolution in SBPs by combining a set of biochemical, biophysical, and structural studies on the SBPs involved in translocation of substrates across the membrane using ATP-binding cassette (ABC) transporters and tripartite ATP-independent periplasmic (TRAP) transporters in gram-negative bacteria Thermotoga maritima and Pseudomonas aeruginosa, respectively. Additionally, experimental, and computational methods were used in conjunction to highlight the variation in the dynamics of these SBPs. The results from this study highlight an intricate role of dynamics in complementing the structural alterations that are required for high-affinity ligand binding. Moreover, first ever neutron structure of a SBP was determined during my study to delineate the extensive network of water in the binding cavities of the SBPs that help stabilize larger substrates by forming water-mediated hydrogen bond interactions with the bound substrates. Furthermore, structures of two SBPs from T. maritima were determined in both substrate-free (apo) and substrate-bound (holo) forms and subsequently used for computational molecular dynamics simulation to determine the variation in dynamics due to substrate-binding. The novel TRAP SBP identified in P. aeruginosa was identified as a promiscuous binder of several tricarboxylic acid cycle (TCA) cycle intermediates. A total of six SBP structures were determined using X-ray crystallography and one SBP structure was determined using neutron crystallography. Finally, experimental neutron scattering was used to experimentally characterize the picosecond to nanosecond dynamics in SBPs and highlighted differences in the translational, rotational, and internal dynamical signatures of two SBP isoforms. Overall, the findings of this study can be broadly applied in biotechnology and biosensor development by artificially engineer affinity or specificity for a particular ligand

On the dynamical incompleteness of the Protein Data Bank

Major scientific challenges that are beyond the capability of individuals need to be addressed by multi-disciplinary and multi-institutional consortia. Examples of these endeavours include the Human Genome Project, and more recently, the Structural Genomics (SG) initiative.The SG initiative pursues the expansion of structural coverage to include at least one structural representative for each protein family to derive the remaining structures using homology modelling. However, biological function is inherently connected with protein dynamics that can be studied by knowing different structures of the same protein. This ensemble of structures provides snapshots of protein conformational diversity under native conditions. Thus, sequence redundancy in the Protein Data Bank (PDB) (i.e. crystallization of the same protein under different conditions) is therefore an essential input contributing to experimentally based studies of protein dynamics and providing insights into protein function.In this work, we show that sequence redundancy, a key concept for exploring protein dynamics, is highly biased and fundamentally incomplete in the PDB. Additionally, our results show that dynamical behaviour of proteins cannot be inferred using homologous proteins. Minor to moderate changes in sequence can produce great differences in dynamical behaviour.Nonetheless, the structural and dynamical incompleteness of the PDB is apparently unrelated concepts in SG. While the first could be reversed by promoting the extension of the structural coverage, we would like to emphasize that furtherfocused efforts will be needed to amend the incompleteness of the PDB in terms of dynamical information content, essential to fully understand protein function.Fil: Monzón, Alexander. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Marino Buslje, Cristina. Universidad Nacional de Quilmes; ArgentinaFil: Zea, Diego Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Fornasari, Maria Silvina. Universidad Nacional de Quilmes; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Parisi, Gustavo Daniel. Universidad Nacional de Quilmes; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin