A new version of the coarse-grained (CG) SIRAH force field for proteins has been developed. Modifications to bonded and non-bonded interactions on the existing molecular topologies significantly ameliorate the structural description and flexibility of a non-redundant set of proteins. The SIRAH 2.0 force field has also been ported to the popular simulation package AMBER, which along with the former implementation in GROMACS expands significantly the potential range of users and performance of this CG force field on CPU/GPU codes. As a non-trivial example of application, we undertook the structural and dynamical analysis of the most abundant and conserved calcium-binding protein, namely, Calmodulin (CaM). CaM is constituted by two calcium-binding motifs called EF-hands, which in presence of Calcium specifically recognize a cognate peptide by embracing it. CG simulations of CaM bound to four Calcium ions in the presence or absence of a binding peptide (holo and apo forms, respectively), resulted in good and stable ion coordination. The simulation of the holo form starting from an experimental structure sampled nearnative conformations, retrieving quasi-atomistic precision. Removing the binding peptide enabled the EF-hands to perform large reciprocal movements, comparable to those observed in NMR structures. On the other hand, the isolated peptide starting from the helical conformation experienced spontaneous unfolding, in agreement with previous experimental data. However, repositioning the peptide in the neighborhood of one EF-hand not only prevented the peptide unfolding but also drove CaM to a fully bound conformation with both EF-hands embracing the cognate peptide, resembling the experimental holo structure. Therefore, SIRAH 2.0 showed the capacity to handle a number of structurally and dynamically challenging situations including metal ion coordination, unbiased conformational sampling, and specific protein-peptide recognition.

BackgroundInfluenza A virus (IAV) is a member of the family Orthomyxoviridae and contains eight segments of a single-stranded RNA genome with negative polarity. The first influenza pandemic of this century was declared in April of 2009, with the emergence of a novel H1N1 IAV strain (H1N1pdm) in Mexico and USA. Understanding the extent and causes of biases in codon usage is essential to the understanding of viral evolution. A comprehensive study to investigate the effect of selection pressure imposed by the human host on the codon usage of an emerging, pandemic IAV strain and the trends in viral codon usage involved over the pandemic time period is much needed.ResultsWe performed a comprehensive codon usage analysis of 310 IAV strains from the pandemic of 2009. Highly biased codon usage for Ala, Arg, Pro, Thr and Ser were found. Codon usage is strongly influenced by underlying biases in base composition. When correspondence analysis (COA) on relative synonymous codon usage (RSCU) is applied, the distribution of IAV ORFs in the plane defined by the first two major dimensional factors showed that different strains are located at different places, suggesting that IAV codon usage also reflects an evolutionary process.ConclusionsA general association between codon usage bias, base composition and poor adaptation of the virus to the respective host tRNA pool, suggests that mutational pressure is the main force shaping H1N1 pdm IAV codon usage. A dynamic process is observed in the variation of codon usage of the strains enrolled in these studies. These results suggest a balance of mutational bias and natural selection, which allow the virus to explore and re-adapt its codon usage to different environments. Recoding of IAV taking into account codon bias, base composition and adaptation to host tRNA may provide important clues to develop new and appropriate vaccines.

A new version of the coarse-grained (CG) SIRAH force field for proteins has been developed. Modifications to bonded and non-bonded interactions on the existing molecular topologies significantly ameliorate the structural description and flexibility of a non-redundant set of proteins. The SIRAH 2.0 force field has also been ported to the popular simulation package AMBER, which along with the former implementation in GROMACS expands significantly the potential range of users and performance of this CG force field on CPU/GPU codes. As a non-trivial example of application, we undertook the structural and dynamical analysis of the most abundant and conserved calcium-binding protein, namely, Calmodulin (CaM). CaM is constituted by two calcium-binding motifs called EF-hands, which in presence of Calcium specifically recognize a cognate peptide by embracing it. CG simulations of CaM bound to four Calcium ions in the presence or absence of a binding peptide (holo and apo forms, respectively), resulted in good and stable ion coordination. The simulation of the holo form starting from an experimental structure sampled nearnative conformations, retrieving quasi-atomistic precision. Removing the binding peptide enabled the EF-hands to perform large reciprocal movements, comparable to those observed in NMR structures. On the other hand, the isolated peptide starting from the helical conformation experienced spontaneous unfolding, in agreement with previous experimental data. However, repositioning the peptide in the neighborhood of one EF-hand not only prevented the peptide unfolding but also drove CaM to a fully bound conformation with both EF-hands embracing the cognate peptide, resembling the experimental holo structure. Therefore, SIRAH 2.0 showed the capacity to handle a number of structurally and dynamically challenging situations including metal ion coordination, unbiased conformational sampling, and specific protein-peptide recognition. 1-28TOC. 2-28 METHODS Re-parameterization of the Model.SIRAH uses a classical Hamiltonian common to most all-atoms potentials to describe particle-particle interactions. Briefly, bonds and angles were described by harmonic terms (e.g.: k b /2*(r-r b ) 2 , with force constant k b and equilibrium values r b ), while Fourier expansions were used for dihedrals. Non-bonded contributions were accounted by Coulomb and Particle Mesh Ewald summation (PME 34,35 ), while Lennard-Jones (LJ) terms were calculated using the standard 12-6 expression (i.e.: 4**[(/r) 12 -(/r) 6 ]). Lorentz-Berthelot combination rules were used for most atom-types pairs. However, specific LJ parameters were set for tuning interactions between some bead pairs (see below). The AMBER scaling factors SCEE=1.2 and SCNB=2.0 were used for the 1-4 non-bonded interactions.3-28 7-28 22-28

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Hepatitis C Virus (HCV) infection treatment has dramatically changed with the advent of direct-acting antiviral agents (DAAs). However, the efficacy of DAAs can be attenuated by the presence of resistance-associated substitutions (RASs) before and after treatment. Indeed, RASs detected in DAA treatment-naïve HCV-infected patients could be useful for clinical management and outcome prediction. Although the frequency of naturally occurring HCV NS5A and NS5B RASs has been addressed in many countries, there are only a few reports on their prevalence in the South American region. The aim of this study was to investigate the presence of RASs to NS5A and NS5B inhibitors in a DAA treatment naïve cohort of Uruguayan patients infected with chronic hepatitis C and compare them with reports from other South American countries. Here, we found that naturally occurring substitutions conferring resistance to NS5A and NS5B inhibitors were present in 8% and 19.2%, respectively, of treatment-naïve HCV genotype 1 infected patients. Importantly, the baseline substitutions in NS5A and NS5B herein identified differ from the studies previously reported in Brazil. Furthermore, Uruguayan strains subtype 1a clustered within all major world clades, showing that HCV variants currently circulating in this country are characterized by a remarkable genetic diversity.

Zika virus (ZIKV) is a member of the family Flaviviridae. In 2015, ZIKV triggered an epidemic in Brazil and spread across Latin America. By May of 2016, the World Health Organization warns over spread of ZIKV beyond this region. Detailed studies on the mode of evolution of ZIKV strains are extremely important for our understanding of the emergence and spread of ZIKV populations. In order to gain insight into these matters, a Bayesian coalescent Markov Chain Monte Carlo analysis of complete genome sequences of recently isolated ZIKV strains was performed. The results of these studies revealed a mean rate of evolution of 1.20 × 10(-3) nucleotide substitutions per site per year (s/s/y) for ZIKV strains enrolled in this study. Several variants isolated in China are grouped together with all strains isolated in Latin America. Another genetic group composed exclusively by Chinese strains were also observed, suggesting the co-circulation of different genetic lineages in China. These findings indicate a high level of diversification of ZIKV populations. Strains isolated from microcephaly cases do not share amino acid substitutions, suggesting that other factors besides viral genetic differences may play a role for the proposed pathogenesis caused by ZIKV infection. J. Med. Virol. 88:1672-1676, 2016. © 2016 Wiley Periodicals, Inc.