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Haptoglobin related protein (Hpr) is a component of the trypanosome lytic factor (TLF), a complex acting in the innate immune response against African trypanosomes. Like haptoglobin (Hp), Hpr binds hemoglobin (Hb) in the blood, but unlike Hp, Hpr does not bind the CD163 receptor. Moreover, unlike Hp, Hpr retains the

Haptoglobin (Hp) counterbalances the adverse effects of extra-erythrocytic hemoglobin (Hb) by trapping the alpha beta dimers of Hb in the bloodstream. In turn, the Hp:Hb complexes display Hb-like reactivity. Here, the kinetics of NO dissociation from ferrous nitrosylated Hp:Hb complexes (i.e., Hp1-1:Hb(II)-NO and Hp2-2:Hb(II)-NO, respectively) are reported at pH 7.0 and 20.0 degrees C. NO dissociation from Hp:Hb(II)-NO complexes has been followed by replacing NO with CO. Denitrosylation kinetics of Hp1-1:Hb(II)-NO and Hp2-2:Hb(II)-NO are biphasic, the relative amplitude of the fast and slow phase being 0.495 +/- 0.015 and 0.485 +/- 0.025, respectively. Values of k(off(NO)1) and k(off(NO)2) (i.e., (6.4 +/- 0.8) x 10(-5) s(-1) and (3.6 +/- 0.6) x 10(-5) s(-1) for Hp1-1:Hb(II)-NO and (5.8 +/- 0.8) x 10(-5) s(-1) and (3.1 +/- 0.6) x 10(-5) s(-1) for Hp2-2:Hb(II)-NO) are unaffected by allosteric effectors and correspond to those reported for the alpha and beta subunits of tetrameric Hb(II)-NO and isolated alpha(II)-NO and beta(II)-NO chains, respectively. This highlights the view that the conformation of the Hb alpha(1)beta(1) and alpha(2)beta(2) dimers matches that of the Hb high affinity conformation. Moreover, the observed functional heterogeneity reflects the variation of energy barriers for the ligand detachment and exit pathway(s) associated to the different structural arrangement of the two subunits in the nitrosylated R-state. Noteworthy, the extent of the inequivalence of alpha and beta chains is closely similar for the O-2, NO and CO dissociation in the R-state, suggesting that it is solely determined by the structural difference between the two subunits.

Butylated hydroxytoluene (BHT) is one of the most commonly used synthetic antioxidants in food, cosmetic, pharmaceutical and petrochemical products. BHT is considered safe for human health, however, its widespread use together with the potential toxicological effects have increased consumers concern about the use of this synthetic food additive. In addition, the estimated daily intake of BHT has been demonstrated to exceed the recommended acceptable threshold. In the present work, using BHT as a case study, the usefulness of computational techniques, such as reverse screening and molecular docking, in identifying protein&ndash, ligand interactions of food additives at the bases of their toxicological effects has been probed. The computational methods here employed have been useful for the identification of several potential unknown targets of BHT, suggesting a possible explanation for its toxic effects. In silico analyses can be employed to identify new macromolecular targets of synthetic food additives and to explore their functional mechanisms or side effects. Noteworthy, this could be important for the cases in which there is an evident lack of experimental studies, as is the case for BHT.

Haptoglobin (Hp) counterbalances the adverse effects of extra-erythrocytic hemoglobin (Hb) by trapping the αβ dimers of Hb in the bloodstream. In turn, the Hp:Hb complexes display Hb-like reactivity. Here, the kinetics of NO dissociation from ferrous nitrosylated Hp:Hb complexes (i.e., Hp1-1:Hb(II)-NO and Hp2-2:Hb(II)-NO, respectively) are reported at pH 7.0 and 20.0 °C. NO dissociation from Hp:Hb(II)-NO complexes has been followed by replacing NO with CO. Denitrosylation kinetics of Hp1-1:Hb(II)-NO and Hp2-2:Hb(II)-NO are biphasic, the relative amplitude of the fast and slow phase being 0.495 ± 0.015 and 0.485 ± 0.025, respectively. Values of k

Pseudomonas aeruginosa is an opportunistic human pathogen highly relevant from a biomedical viewpoint. It is one of the main causes of infection in hospitalized patients and a major cause of mortality of cystic fibrosis patients. This is also due to its ability to develop resistance to antibiotics by various mechanisms. Therefore, it is urgent and desirable to identify novel targets for the development of new antibacterial drugs against Pseudomonas aeruginosa. In this work this problem was tackled by an in silico approach aimed at providing a reliable structural model and functional annotation for the Pseudomonas aeruginosa periplasmic proteins for which these data are not available yet. A total of 83 protein sequences were analyzed, and the corresponding structural models were built, leading to the identification of 32 periplasmic ‘substrate-binding proteins’, 14 enzymes and 4 proteins with different functions, including lipids and metals binding. The most interesting cases were found within the ‘enzymes’ group with the identification of a lipase, which can be regarded as a virulence factor, a protease involved in the assembly of β-barrel membrane proteins and a l,d-transpeptidase, which could contribute to confer resistance to β-lactam antibiotics to the bacterium. Communicated by Ramaswamy H. Sarma.

Members of the major facilitator superfamily of transporters (MFS) play an essential role in many physiological processes such as development, neurotransmission, and signaling. Aberrant functions of MFS proteins are associated with several diseases, including cancer, schizophrenia, epilepsy, amyotrophic lateral sclerosis and Alzheimer&rsquo, s disease. MFS transporters are also involved in multidrug resistance in bacteria and fungi. The structures of most MFS members, especially those of members with significant physiological relevance, are yet to be solved. The lack of structural and functional information impedes our detailed understanding, and thus the pharmacological targeting, of these transporters. To improve our knowledge on the mechanistic principles governing the function of MSF members, molecular dynamics (MD) simulations were performed on the inward-facing and outward-facing crystal structures of the human ferroportin homologue from the Gram-negative bacterium Bdellovibrio bacteriovorus (BdFpn). Several simulations with an excess of iron ions were also performed to explore the relationship between the protein&rsquo, s dynamics and the ligand recognition mechanism. The results reinforce the existence of the alternating-access mechanism already described for other MFS members. In addition, the reorganization of salt bridges, some of which are conserved in several MFS members, appears to be a key molecular event facilitating the conformational change of the transporter.

Motivation: Bringing a new drug to the market is expensive and time-consuming. To cut the costs and time, computer-aided drug design (CADD) approaches have been increasingly included in the drug discovery pipeline. However, despite traditional docking tools show a good conformational space sampling ability, they are still unable to produce accurate binding affinity predictions. This work presents a novel scoring function for molecular docking seamlessly integrated into DockingApp, a user-friendly graphical interface for AutoDock Vina. The proposed function is based on a random forest model and a selection of specific features to overcome the existing limits of Vina&rsquo, s original scoring mechanism. A novel version of DockingApp, named DockingApp RF, has been developed to host the proposed scoring function and to automatize the rescoring procedure of the output of AutoDock Vina, even to nonexpert users. Results: By coupling intermolecular interaction, solvent accessible surface area features and Vina&rsquo, s energy terms, DockingApp RF&rsquo, s new scoring function is able to improve the binding affinity prediction of AutoDock Vina. Furthermore, comparison tests carried out on the CASF-2013 and CASF-2016 datasets demonstrate that DockingApp RF&rsquo, s performance is comparable to other state-of-the-art machine-learning- and deep-learning-based scoring functions. The new scoring function thus represents a significant advancement in terms of the reliability and effectiveness of docking compared to AutoDock Vina&rsquo, s scoring function. At the same time, the characteristics that made DockingApp appealing to a wide range of users are retained in this new version and have been complemented with additional features.

Haptoglobin related protein (Hpr) is a component of the trypanosome lytic factor (TLF), a complex acting in the innate immune response against African trypanosomes. Like haptoglobin (Hp), Hpr binds hemoglobin (Hb) in the blood, but unlike Hp, Hpr does not bind the CD163 receptor. Moreover, unlike Hp, Hpr retains the N-terminal signal peptide that is required for the association with Apolipoprotein L-1 (ApoL-1), a component of the TLF complex. Here, the molecular model of human Hpr has been built based on the high sequence identity with human Hp (91%). The structural bases of Hpr:Hpr dimerization and Hpr recognition by Hb and Trypanosoma brucei brucei Hp receptor (TbHpHbR) have been analyzed in parallel with those of Hp:Hp, Hp:Hb, and TbHpHbR:Hp:Hb complexes. We show that the Cys33-Cys33 intermolecular disulfide bridge that stabilizes the Hp1:Hp1 complex is replaced by the Phe33, Pro34, and Phe48 hydrophobic core in the Hpr:Hpr dimer. Moreover, we show that the N-terminal peptide of Hpr participates in the stabilization of the Hpr:Hpr dimer. Thus, the N-terminal peptide seems to have been retained in Hpr to mediate its critical role in the human innate immunity towards Trypanosoma brucei brucei infection. Communicated by Ramaswamy H. Sarma

Blood-feeding animals are known for their ability to produce bioactive compounds to impair haemostasis and suppress pain perception in the host. These compounds are extremely appealing for pharmacological development since they are generally very effective and specific for their molecular target. A preliminary RNA-Seq based characterization of the secretion from salivary and mid-oesophageal tissues of the vampire snail Cumia reticulata, revealed a complex mixture of feeding-related transcripts with potential anaesthetic and anticoagulant action. Based on the cloned full-length mRNAs, it was possible to verify the sequence of five genes encoding haematophagy-related products. The in silico modelled three-dimensional structure of each translational product was analysed to gain information on their potential biochemical activity. We have hereby validated and further investigated the assembled transcripts presumably involved in the antihaemostatic action, to improve our comprehensive understanding of this subset of the feeding secretion. The studied proteins included both inhibitors of primary haemostasis such as the vWFA domain-containing proteins, and compounds targeting different steps of the coagulation cascade, as e.g. the Turripeptide-like/protease inhibitor, the TFPI-like multiple Kunitz-type protease inhibitors, the Meprin-like metalloproteases and the Astacin/ShKT-like domain-containing proteins. All these molecules showed promising potential for pharmacological development.

In recent years, the high throughput and the low cost of next-generation sequencing (NGS) technologies have led to an increase of the amount of (meta)genomic data, revolutionizing genomic research studies. However, the quality of sequencing data could be affected by experimental errors derived from defective methods and protocols. This represents a serious problem for the scientific community with a negative impact on the correctness of studies that involve genomic sequence analysis. As a countermeasure, several alignment and taxonomic classification tools have been developed to uncover and correct errors. In this critical review some of these integrated software tools and pipelines used to detect contaminations in reference genome databases and sequenced samples are reported. In particular, case studies of bacterial contaminations, contaminations of human origin, mitochondrial contaminations of ancient DNA, and cross contaminations are examined.