The purpose of this quick guide is to help new modelers who have little or no background in comparative modeling yet are keen to produce high-resolution protein 3D structures for their study by following systematic good modeling practices, using affordable personal computers or online computational resources. Through the available experimental 3D-structure repositories, the modeler should be able to access and use the atomic coordinates for building homology models. We also aim to provide the modeler with a rationale behind making a simple list of atomic coordinates suitable for computational analysis abiding to principles of physics (e.g., molecular mechanics). Keeping that objective in mind, these quick tips cover the process of homology modeling and some postmodeling computations such as molecular docking and molecular dynamics (MD). A brief section was left for modeling nonprotein molecules, and a short case study of homology modeling is discussed.

• MeSH
• algoritmy MeSH
• aminokyseliny chemie   MeSH
• biologické modely MeSH
• databáze proteinů MeSH
• internet MeSH
• ionty MeSH
• koncentrace vodíkových iontů MeSH
• ligandy MeSH
• počítačová simulace MeSH
• posttranslační úpravy proteinů MeSH
• proteiny chemie   MeSH
• rozpouštědla MeSH
• sbalování proteinů MeSH
• simulace molekulární dynamiky MeSH
• simulace molekulového dockingu MeSH
• software MeSH
• strojové učení MeSH
• strukturní homologie proteinů MeSH
• voda MeSH
• výpočetní biologie metody   MeSH
• zobrazování trojrozměrné metody   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The human dopamine, norepinephrine, and serotonin transporters (hDAT, hNET, and hSERT) are carriers of neurotransmitters and targets for many drugs. Pioneering works in the past three years to elucidate experimental models of the Drosophila dDAT and human hSERT structures will rapidly impact the field of neuroscience. Here, we evaluated automated homology-based human models of these transporters, employing systematic physics-based, knowledge-based, and empirical-based check. Modeling guidelines were conveyed with attention to the central binding site (S1), secondary binding site (S2), and the extracellular loops EL2 and EL4. Application of new experimental models (dDAT and hSERT) will improve the accuracy of homology models, previously utilizing prokaryotic leucine transporter (LeuT) structure, and provide better predictions of ligand interactions, which is required for understanding of cellular mechanisms and for development of novel therapeutics.

• MeSH
• acetyltransferasy genetika   metabolismus   MeSH
• Drosophila MeSH
• konformace proteinů MeSH
• lidé MeSH
• membránové transportní proteiny pro serotonin genetika   metabolismus   MeSH
• molekulární modely * MeSH
• proteiny Drosophily genetika   metabolismus   MeSH
• proteiny přenášející dopamin přes plazmatickou membránu genetika   metabolismus   MeSH
• proteiny přenášející noradrenalin přes plazmatickou membránu genetika   metabolismus   MeSH
• rozpoznávání automatizované MeSH
• sekvenční homologie aminokyselin * MeSH
• vazebná místa MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH

Twelve homology models of the human M2 muscarinic receptor using different sets of templates have been designed using the Prime program or the modeller program and compared to crystallographic structure (PDB:3UON). The best models were obtained using single template of the closest published structure, the M3 muscarinic receptor (PDB:4DAJ). Adding more (structurally distant) templates led to worse models. Data document a key role of the template in homology modeling. The models differ substantially. The quality checks built into the programs do not correlate with the RMSDs to the crystallographic structure and cannot be used to select the best model. Re-docking of the antagonists present in crystallographic structure and relative binding energy estimation by calculating MM/GBSA in Prime and the binding energy function in YASARA suggested it could be possible to evaluate the quality of the orthosteric binding site based on the prediction of relative binding energies. Although estimation of relative binding energies distinguishes between relatively good and bad models it does not indicate the best one. On the other hand, visual inspection of the models for known features and knowledge-based analysis of the intramolecular interactions allows an experimenter to select overall best models manually.

• MeSH
• konformace proteinů * MeSH
• krystalografie rentgenová * MeSH
• lidé MeSH
• molekulární modely MeSH
• receptor muskarinový M2 chemie   MeSH
• sekvence aminokyselin MeSH
• sekvenční homologie aminokyselin MeSH
• simulace molekulového dockingu MeSH
• vazebná místa MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

To find an effective drug for Zika virus, it is important to understand how numerous proteins which are critical for the virus' structure and function interact with their counterparts. One approach to inhibiting the flavivirus is to deter its ability to bind onto glycoproteins; however, the crystal structures of envelope proteins of the ever-evolving viral strains that decipher glycosidic or drug-molecular interactions are not always available. To fill this gap, we are reporting a holistic, simulation-based approach to predict compounds that will inhibit ligand binding onto a structurally unresolved protein, in this case the Zika virus envelope protein (ZVEP), by developing a three-dimensional general structure and analyzing sites at which ligands and small drug-like molecules interact. By examining how glycan molecules and small-molecule probes interact with a freshly resolved ZVEP homology model, we report the susceptibility of ZVEP to inhibition via two small molecules, ZINC33683341 and ZINC49605556-by preferentially binding onto the primary receptor responsible for the virus' virulence. Antiviral activity was confirmed when ZINC33683341 was tested in cell culture. We anticipate the results to be a starting point for drug discovery targeting Zika virus and other emerging pathogens.

• MeSH
• antivirové látky chemie   farmakologie   MeSH
• Cercopithecus aethiops MeSH
• knihovny malých molekul chemie   farmakologie   MeSH
• molekulární modely MeSH
• počítačová simulace MeSH
• polysacharidy metabolismus   MeSH
• proteiny virového obalu antagonisté a inhibitory   chemie   MeSH
• strukturní homologie proteinů MeSH
• vazebná místa MeSH
• Vero buňky MeSH
• virová nálož účinky léků   MeSH
• virus zika účinky léků   metabolismus   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Secreted aspartic proteases (Saps) of pathogenic Candida spp. represent a specific target for antifungal drug development. We synthesized a series of peptidomimetic inhibitors with different isosteric groups and modifications at individual positions and tested them with purified Saps from C. albicans (Sap2p), C. tropicalis (Sapt1p), and C. parapsilosis (Sapp1p). The kinetic parameters indicated that all three proteases prefer binding of inhibitors containing bulky hydrophobic residues between positions P3 and P3'. The most divergent specificity was found for Sapp1p. The sequence alignment of Sap2p, Sapt1p, and Sapp1p, and homology modeling of Sapp1p with the crystal structure of Sapt1p and the complex of Sap2p with a peptidomimetic inhibitor showed that the overall folds of Sap2p, Sapt1p, and Sapp1p are similar. However, the N- and C-terminal loops formed by disulfide bonds between residues 47-53 and 258-292 are significantly shorter in Sapp1p, and a unique insertion following Tyr 129 in Sapp1p results in the formation of a loop that can interact with inhibitor residues. These Sapp1p structural differences might lead to its altered susceptibility to inhibition.

• MeSH
• aspartátové endopeptidasy chemie   účinky léků   MeSH
• Candida enzymologie   MeSH
• financování organizované MeSH
• inhibitory proteas farmakologie   chemická syntéza   chemie   MeSH
• izoenzymy chemie   účinky léků   MeSH
• molekulární konformace MeSH
• molekulární modely MeSH
• molekulární sekvence - údaje MeSH
• sekvence aminokyselin MeSH
• sekvenční homologie aminokyselin MeSH
• stereoizomerie MeSH
• substrátová specifita MeSH
• terciární struktura proteinů MeSH
• vztahy mezi strukturou a aktivitou MeSH

β-N-acetylglucosaminidases from the family 84 of glycoside hydrolases form a small group of glycosidases in eukaryotes responsible for the modification of nuclear and cytosolic proteins with O-GlcNAc, thus they are involved in a number of important cell processes. Here, the first fungal β-N-acetylglucosaminidase from Penicillium chrysogenum was expressed in Pichia pastoris and secreted into the media, purified and characterized. Moreover, homology modeling and substrate and inhibitor docking were performed to obtain structural information on this new member of the GH84 family. Surprisingly, we found that this fungal β-N-acetylglucosaminidase with its sequence and structure perfectly fitting to the GH84 family displays biochemical properties rather resembling the β-N-acetylhexosaminidases from the family 20 of glycoside hydrolases. This work helped to increase the knowledge on the scarcely studied glycosidase family and revealed a new type of eukaryotic β-N-acetylglucosaminidase.

• MeSH
• acetylglukosaminidasa chemie   genetika   izolace a purifikace   metabolismus   MeSH
• molekulární sekvence - údaje MeSH
• Penicillium chrysogenum enzymologie   genetika   MeSH
• Pichia genetika   metabolismus   MeSH
• rekombinantní proteiny chemie   genetika   izolace a purifikace   metabolismus   MeSH
• sekvence aminokyselin MeSH
• sekvenční seřazení MeSH
• simulace molekulového dockingu MeSH
• vazba proteinů MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Targeted therapy is a promising approach for treatment of neuroblastoma as evident from the large number of targeting agents employed in clinical practice today. In the absence of known crystal structures, researchers rely on homology modeling to construct template-based theoretical structures for drug design and testing. Here, we discuss three candidate cell surface proteins that are suitable for homology modeling: human norepinephrine transporter (hNET), anaplastic lymphoma kinase (ALK), and neurotrophic tyrosine kinase receptor 2 (NTRK2 or TrkB). When choosing templates, both sequence identity and structure quality are important for homology modeling and pose the first of many challenges in the modeling process. Homology modeling of hNET can be improved using template models of dopamine and serotonin transporters instead of the leucine transporter (LeuT). The extracellular domains of ALK and TrkB are yet to be exploited by homology modeling. There are several idiosyncrasies that require direct attention throughout the process of model construction, evaluation and refinement. Shifts/gaps in the alignment between the template and target, backbone outliers and side-chain rotamer outliers are among the main sources of physical errors in the structures. Low-conserved regions can be refined with loop modeling method. Residue hydrophobicity, accessibility to bound metals or glycosylation can aid in model refinement. We recommend resolving these idiosyncrasies as part of "good modeling practice" to obtain highest quality model. Decreasing physical errors in protein structures plays major role in the development of targeting agents and understanding of chemical interactions at the molecular level.

• Publikační typ
• časopisecké články MeSH

Fungal β-N-acetylhexosaminidases are inducible extracellular enzymes with many biotechnological applications. The enzyme from Penicillium oxalicum has unique enzymatic properties despite its close evolutionary relationship with other fungal hexosaminidases. It has high GalNAcase activity, tolerates substrates with the modified N-acyl group better and has some other unusual catalytic properties. In order to understand these features, we performed isolation, biochemical and enzymological characterization, molecular cloning and molecular modelling. The native enzyme is composed of two catalytic units (65 kDa each) and two propeptides (15 kDa each), yielding a molecular weight of 160 kDa. Enzyme deglycosylated by endoglycosidase H had comparable activity, but reduced stability. We have cloned and sequenced the gene coding for the entire hexosaminidase from P. oxalicum. Sufficient sequence identity of this hexosaminidase with the structurally solved enzymes from bacteria and humans with complete conservation of all catalytic residues allowed us to construct a molecular model of the enzyme. Results from molecular dynamics simulations and substrate docking supported the experimental kinetic and substrate specificity data and provided a molecular explanation for why the hexosaminidase from P. oxalicum is unique among the family of fungal hexosaminidases.

• MeSH
• beta-N-acetylhexosaminidasy chemie   genetika   izolace a purifikace   metabolismus   MeSH
• fungální proteiny chemie   genetika   izolace a purifikace   metabolismus   MeSH
• glykosylace MeSH
• katalytická doména MeSH
• kinetika MeSH
• koncentrace vodíkových iontů MeSH
• konzervovaná sekvence MeSH
• mannosyl-glykoprotein endo-beta-N-acetylglukosaminidasa metabolismus   MeSH
• molekulární modely MeSH
• molekulární sekvence - údaje MeSH
• Penicillium enzymologie   genetika   MeSH
• prekurzory enzymů chemie   genetika   izolace a purifikace   metabolismus   MeSH
• sekundární struktura proteinů MeSH
• sekvence aminokyselin MeSH
• sekvenční homologie aminokyselin MeSH
• sekvenční seřazení MeSH
• simulace molekulární dynamiky MeSH
• stabilita enzymů MeSH
• substrátová specifita MeSH
• teplota MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

Trojrozměrné struktury proteinů je možné předpovídat tak, že vezmeme denaturovaný protein a na počítači simulujeme proces jeho sbalení. Tento článek shrnuje úspěchy i úskalí tohoto postupu, zejména využití vysoce výkonných počítačů, projektů distribuovaných výpočtů, grafických karet a specializovaných počítačů.

The three-dimensional structure of a protein can be predicted by a simulation of its folding from fully denaturated state. This article review success stories as well as pitfalls of this approach, namely applications of high performance computers, distributed computing projects, graphical processing units and specialised hardware.

• MeSH
• počítačová simulace * MeSH
• sbalování proteinů * MeSH
• strukturní homologie proteinů MeSH

Lipid-solubilní koenzym Q, který je syntetizován ve všech tkáních organizmu a zasahuje do celé řady buněčných dějů, má specifické postavení mezi antioxidanty. Cílem současné studie bylo srovnat antioxidační působení některých homologů a analogů koenzymu Q na stejném modelovém systému membrán mozkové kůry potkana in vitro. Zjistili jsme, že všechny použité látky zabraňovaly v různé míře vzniku lipidových peroxidací. Nejúčinnější byl koenzym Q2. Výsledky ukazují, že některé homology a analogy koenzymu Q mohou sloužit (podobně jako idebenon) k vývoji látek pro léčbu nemocí, na kterých se podílí oxidační stres.

A lipid-soluble coenzyme Q, which is synthesized in tissues and is involved in a variety of cellular processes, has a specific place among antioxidants. The aim of our study was to compare the protective effects of some homologues and analogues of coenzyme Q on the same model system of cerebral cortex membranes in vitro against damage of lipid peroxidation. Our results revealed that used substances prevented the increase in the formation of thiobarbituric acid-reactive substances. Coenzyme Q2 was the most efficient in preventing lipid peroxidation. Our results show that some analogues and homologues of coenzyme Q could serve (similarly as idebenone) for the development of pharmaceutical compounds for treatment of diseases in which oxidative stress is present.

• MeSH
• antioxidancia MeSH
• buněčná membrána chemie   MeSH
• finanční podpora výzkumu jako téma MeSH
• látky reagující s kyselinou thiobarbiturovou MeSH
• mozková kůra MeSH
• oxidační stres MeSH
• peroxidace lipidů účinky záření   MeSH
• techniky in vitro MeSH
• ubichinon analogy a deriváty   farmakologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH