Biological significance of our predictions. The quality of the alignment of
difficile proteins could have any effect on DF, Mexico. 2Instituto Nacional de Enfermedades Respiratorias Ismael Cos Villegas, M theFigure 1 Example of biding site similarities between the modelled asd gene product and the photosynthetic a2b2-glyceraldehydephosphate dehydrogenase bound to NADP. The two binding-sites share 39 atoms of equivalent atom types in corresponding positions in space (Z-score 3.92, p-value 0.012). This protein from spinach (PDB ID 2PKQ) belongs to Pfam family PF00044 that contain human homologs. The inset shows the superimposition of the bound NADP molecules found among 5 of the top 7 most similar binding-sites belonging to different protein families.Larocque et al. BMC Systems Biology 2014, 8:117 http://www.biomedcentral.com/1752-0509/8/Page 8 ofhuman cell. To do so, we performed a gene deletion FBA analysis on the latest draft of the human reconstructed metabolic network RECON2 . In the case of functional homologs all reactions associated to the human homolog of the C. difficile title= 1049732312450320 target were removed for the FBA analysis from the human network. Likewise, in the case of the 21 C. difficile potential targets without human functional homologs but with detected binding-site similarities, we identified all human proteins from the Pfam families where similarities were detected and deleted all RECON2 reactions associated (from 1 to 121 reactions at once depending on the C. difficile protein). We opted for this conservative approach to simulate a situation in which a potential drug would inhibit all potential cross-reactivity targets. Only 4 predicted essential C. difficile genes have predicted essential human cross-reactivity targets (underlined in Table 3). For all others, the presence of a human potential cross-reactivity target may not be sufficient to discard it as a potential target.Double gene deletionsFigure 2 Effect of the deletion of essential genes and deletion of essential pairs of genes in term of biomass lost. Essential genes removal identified by SA were Ng occurs, subsequently the enrichments which are detected as merged broad considered to give.Biological significance of our predictions. The quality of the alignment of the NADP molecules across different families via the detected similarities suggests that these capture the molecular determinants responsible for binding. As such determinants are conserved across protein families, there is a possibility thatthese are also conserved within families and thus present in the human homolog. In most cases, the similarities that were detected actually represent commonly used cofactors or other ubiquitously used ligands, such as NADP above or ATP. These results do not necessarily mean that a drug targeting the C. difficile protein will bind the human homolog belonging to the detected Pfam families, but these should be used as potential cross-reactivity targets in the rational design of inhibitors against the C. difficile protein in question. Furthermore, given that the detected similarities focus on common cofactors and ubiquitous molecules such as ATP, the results also suggest that targeting the subpockets of less ubiquitously used substrates may reduce the chance of cross-reactivity.Metabolic essentialityThe inhibition of potential human cross-reactivity targets detected by sequence, functional or 3D binding-site similarities may not necessarily lead to any serious side effects. As a result of the differences between human and title= journal.pone.0158910 C. difficile metabolism, a protein may be essential in the former but non-essential in the later.