Structure, substrate binding, and symmetry of the mitochondrial ADP/ATP carrier in its matrix-open state.

The mitochondrial ADP/ATP carrier (AAC) performs the first and last step in oxidative phosphorylation by exchanging ADP and ATP across the mitochondrial inner membrane. Its optimal function has been shown to be dependent on cardiolipins (CLs), unique phospholipids located almost exclusively in the mitochondrial membrane. In addition, AAC exhibits an enthralling threefold pseudosymmetry, a unique feature of members of the SLC25 family. Recently, its conformation poised for binding of ATP was solved by x-ray crystallography referred to as the matrix state. Binding of the substrate leads to conformational changes that export of ATP to the mitochondrial intermembrane space. In this contribution, we investigate the influence of CLs on the structure, substrate-binding properties, and structural symmetry of the matrix state, employing microsecond-scale molecular dynamics simulations. Our findings demonstrate that CLs play a minor stabilizing role on the AAC structure. The interdomain salt bridges and hydrogen bonds forming the cytoplasmic network and tyrosine braces, which ensure the integrity of the global AAC scaffold, highly benefit from the presence of CLs. Under these conditions, the carrier is found to be organized in a more compact structure in its interior, as revealed by analyses of the electrostatic potential, measure of the AAC cavity aperture, and the substrate-binding assays. Introducing a convenient structure-based symmetry metric, we quantified the structural threefold pseudosymmetry of AAC, not only for the crystallographic structure, but also for conformational states of the carrier explored in the molecular dynamics simulations. Our results suggest that CLs moderately contribute to preserve the pseudosymmetric structure of AAC.

Correlation of membrane protein conformational and functional dynamics.

Conformational changes in ion channels lead to gating of an ion-conductive pore. Ion flux has been measured with high temporal resolution by single-channel electrophysiology for decades. However, correlation between functional and conformational dynamics remained difficult, lacking experimental techniques to monitor sub-millisecond conformational changes. Here, we use the outer membrane protein G (OmpG) as a model system where loop-6 opens and closes the ß-barrel pore like a lid in a pH-dependent manner. Functionally, single-channel electrophysiology shows that while closed states are favored at acidic pH and open states are favored at physiological pH, both states coexist and rapidly interchange in all conditions. Using HS-AFM height spectroscopy (HS-AFM-HS), we monitor sub-millisecond loop-6 conformational dynamics, and compare them to the functional dynamics from single-channel recordings, while MD simulations provide atomistic details and energy landscapes of the pH-dependent loop-6 fluctuations. HS-AFM-HS offers new opportunities to analyze conformational dynamics at timescales of domain and loop fluctuations.

A Linear Interaction Energy Model for Cavitand Host-Guest Binding Affinities.

Host-guest systems provide excellent models to explore molecular recognition in solution along with relevant technological applications from drug carriers to chemosensors. Here, we present a linear interaction energy (LIE) model to predict the binding affinity in host-guests with remarkable efficiency and predictive power. Using four host families, including cucurbiturils, octa acids, and ß-cyclodextrin, and a large set (49) of chemically diverse guests, we demonstrate that binding-affinity predictions with a root mean square error <1.5 kcal/mol from experiments can be obtained with a few nanoseconds of molecular dynamics. The parameters of the LIE model are shown to be transferable among host-guest families, and the quality of the predictions is essentially force-field independent. Inclusion of the strain energy of the host in the bound state appears to be critically important to improve the quality of the predictions, particularly when the host and the guest have comparable sizes. Unsuccesful predictions for 28 additional highly charged and bulky guests to cucurbit[7]uril indicate future directions for improvement.

Synthesis and DFT Calculations of Novel Vanillin-Chalcones and Their 3-Aryl-5-(4-(2-(dimethylamino)-ethoxy)-3-methoxyphenyl)-4,5-dihydro-1H-pyrazole-1-carbaldehyde Derivatives as Antifungal Agents.

Novel (E)-1-(aryl)-3-(4-(2-(dimethylamino)ethoxy)-3-methoxyphenyl) prop-2-en-1-ones 4 were synthesized by a Claisen-Schmidt reaction of 4-(2-(dimethylamino)ethoxy)-3-methoxy-benzaldehyde (2) with several acetophenone derivatives 3. Subsequently, cyclocondensation reactions of chalcones 4 with hydrazine hydrate afforded the new racemic 3-aryl-5-(4-(2-(dimethylamino)ethoxy)-3-methoxyphenyl)-4,5-dihydro-1H-pyrazole-1-carbaldehydes 5 when the reaction was carried out in formic acid. The antifungal activity of both series of compounds against eight fungal species was determined. In general, chalcone derivatives 4 showed better activities than pyrazolines 5 against all tested fungi. None of the compounds 4a-g and 5a-g showed activity against the three Aspergillus spp. In contrast, most of the compounds 4 showed moderate to high activities against three dermatophytes (MICs 31.25-62.5 µg/mL), being 4a followed by 4c the most active structures. Interestingly, 4a and 4c possess fungicidal rather than fungistatic activities, with MFC values between 31.25 and 62.5 µg/mL. The comparison of the percentages of inhibition of C. neoformans by the most active compounds 4, allowed us to know the role played by the different substituents of the chalcones' A-ring. Also the most anti-cryptococcal compounds 4a-c and 4g, were tested in a second panel of five clinical C. neoformans strains in order to have an overview of their inhibition capacity not only of standardized but also of clinical C. neoformans strains. DFT calculations showed that the electrophilicity is the main electronic property to explain the differences in antifungal activities for the synthesized chalcones and pyrazolines compounds. Furthermore, a quantitative reactivity analysis showed that electron-withdrawing substituted chalcones presented the higher electrophilic character and hence, the greater antifungal activities among compounds of series 4.

Computational Approaches to the Chemical Equilibrium Constant in Protein-ligand Binding.

The physiological role played by protein-ligand recognition has motivated the development of several computational approaches to the ligand binding affinity. Some of them, termed rigorous, have a strong theoretical foundation but involve too much computation to be generally useful. Some others alleviate the computational burden by introducing strong approximations and/or empirical calibrations, which also limit their general use. Most importantly, there is no straightforward correlation between the predictive power and the level of approximation introduced. Here, we present a general framework for the quantitative interpretation of protein-ligand binding based on statistical mechanics. Within this framework, we re-derive self-consistently the fundamental equations of some popular approaches to the binding constant and pinpoint the inherent approximations. Our analysis represents a first step towards the development of variants with optimum accuracy/efficiency ratio for each stage of the drug discovery pipeline.

2D, 3D-QSAR and molecular docking of 4(1H)-quinolones analogues with antimalarial activities.

Cytochrome bc1 has become a major focus as a molecular target in malaria parasites, which are the most important vector-borne infectious disease in the world. The inhibition of cytochrome bc1 blocks the mitochondrial respiratory chain and the consequent arrest of pyrimidine biosynthesis, which is essential for parasite development. The authors developed a theoretical study of two-dimensional, three-dimensional quantitative structure-activity relationships and a docking analysis of a series of 4(1H)-quinolones acting as cytochrome bc1 inhibitors. The predictive ability of the quantitative structure-activity relationship models was assessed using internal (leave-one-out cross-validation) and external (test set with 8 compounds) validation. From the two-dimensional quantitative structure-activity relationship models, the authors emphasized the following descriptors: GCUT_SLOGP_0, SLogP_VSA_5, Kier molecular flexibility index, electrophilicity index, the partition coefficient and the charge of atom 5 of the quinolone ring as the most important to explain the antimalarial activity of the compounds studied. Three-dimensional quantitative structure-activity relationship models showed that the substituents R1 and R4 in 4(1H)-quinolones analogues are key modulators to enhance the antimalarial activity. The appropriate binding conformations and orientations of these compounds interacting with cytochrome bc1 were also revealed by molecular docking. Based on the established models, 8 new compounds with highly predicted antimalarial activity have been theoretically designed and presented as a reference for synthesis and antimalarial evaluation.