Deep learning of protein sequence design of protein-protein interactions.

MOTIVATION: As more data of experimentally determined protein structures are becoming available, data-driven models to describe protein sequence-structure relationships become more feasible. Within this space, the amino acid sequence design of protein-protein interactions is still a rather challenging subproblem with very low success rates-yet, it is central to most biological processes. RESULTS: We developed an attention-based deep learning model inspired by algorithms used for image-caption assignments to design peptides or protein fragment sequences. Our trained model can be applied for the redesign of natural protein interfaces or the designed protein interaction fragments. Here, we validate the potential by recapitulating naturally occurring protein-protein interactions including antibody-antigen complexes. The designed interfaces accurately capture essential native interactions and have comparable native-like binding affinities in silico. Furthermore, our model does not need a precise backbone location, making it an attractive tool for working with de novo design of protein-protein interactions. AVAILABILITY AND IMPLEMENTATION: The source code of the method is available at https://github.com/strauchlab/iNNterfaceDesign. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Unusual In-plane Aromaticity Facilitates Intramolecular Hydrogen Transfer in Long-Bonded cis-Isonitrosyl Methoxide.

The hydrogen-atom transfer from methoxy radical to nitric oxide, leading to the formation of formaldehyde and nitroxyl, represents a secondary reaction of photodissociation of methyl nitrite, which is used as rocket fuel. In this study, we explored the potential energy profile of the hydrogen-atom transfer using the electronic structure calculations at the DLPNO-CCSD(T)/aug-cc-pVTZ level of theory for two isomeric forms (cis and trans) of the pre-reaction complex. The cis-oriented pre-reaction complex has a weak elongated O─O bond, which gets further elongated in the hydrogen transfer transition state. This O─O bond stabilizes the pre-reaction complex by 32.9 kJ/mol. The O─O-induced stabilization is even greater for the transition state (48.2 kJ/mol), which was unexpected because of the larger O─O distance in the transition state structure. To address this paradox, we performed the electronic structure analysis of the reaction participants using the valence bond (VB) theory, natural resonance theory, topological analysis of the electron density and its derivatives, and analysis of the electron localization function distribution. This combined analysis led to the conclusion that the cis-transition state for hydrogen transfer, instead of being directly stabilized by the O─O interaction, gained substantial stabilization from the in-plane five-center six-electron aromaticity.

Discovery and Characterization of Halogenated Xanthene Inhibitors of DUSP5 as Potential Photodynamic Therapeutics.

Dual specific phosphatases (DUSPs) are an important class of mitogen-activated protein kinase (MAPK) regulators, and are drug targets for treating vascular diseases. Previously we had shown that DUSP5 plays a role in embryonic vertebrate vascular patterning. Herein, we screened a library of FDA-approved drugs and related compounds, using a para-nitrophenylphosphate substrate (pNPP)-based assay. This assay identified merbromin (also known as mercurochrome) as targeting DUSP5; and, we subsequently identified xanthene-ring based merbromin analogs eosin Y, erythrosin B, and rose bengal, all of which inhibit DUSP5 in vitro. Inhibition was time-dependent for merbromin, eosin Y, 2',7'-dibromofluorescein, and 2',7'-dichlorofluorescein, with enzyme inhibition increasing over time. Reaction progress curve data fit best to a slow-binding model of irreversible enzyme inactivation. Potency of the time-dependent compounds, except for 2',7'-dichlorofluorescein, was diminished when dithiothreitol (DTT) was present, suggesting thiol reactivity. Two additional merbromin analogs, erythrosin B and rose bengal also inhibit DUSP5, but have the therapeutic advantage of being less sensitive to DTT and exhibiting little time dependence for inhibition. Inhibition potency is correlated with the xanthene dye's LUMO energy, which affects ability to form light-activated radical anions, a likely active inhibitor form. Consistent with this hypothesis, rose bengal inhibition is light-dependent and demonstrates the expected red shifted spectrum upon binding to DUSP5, with a Kd of 690 nM. These studies provide a mechanistic foundation for further development of xanthene dyes for treating vascular diseases that respond to DUSP5 inhibition, with the following relative potencies: rose bengal > merbromin > erythrosin B > eosin Y.

Role of Conserved Histidine and Serine in the HCXXXXXRS Motif of Human Dual-Specificity Phosphatase 5.

BACKGROUND: The mitogen-activated protein kinase (MAPK) pathway is functionally generic and critical in maintaining physiological homeostasis and normal tissue development. This pathway is under tight regulation, which is in part mediated by dual-specific phosphatases (DUSPs), which dephosphorylate serine, threonine, and tyrosine residues of the ERK family of proteins. DUSP5 is of high clinical interest because of mutations we identified in this protein in patients with vascular anomalies. Unlike other DUSPs, DUSP5 has unique specificity toward substrate pERK1/2. Using molecular docking and simulation strategies, we previously showed that DUSP5 has two pockets, which are utilized in a specific fashion to facilitate specificity toward catalysis of its substrate pERK1/2. Remarkably, most DUSPs share high similarity in their catalytic sites. Studying the catalytic domain of DUSP5 and identifying amino acid residues that are important for dephosphorylating pERK1/2 could be critical in developing small molecules for therapies targeting DUSP5. RESULTS: In this study, we utilized computational modeling to identify and predict the importance of two conserved amino acid residues, H262 and S270, in the DUSP5 catalytic site. Modeling studies predicted that catalytic activity of DUSP5 would be altered if these critical conserved residues were mutated. We next generated independent Glutathione-S-Transferase (GST)-tagged full-length DUSP5 mutant proteins carrying specific mutations H262F and S270A in the phosphatase domain. Biochemical analysis was performed on these purified proteins, and consistent with our computational prediction, we observed altered enzyme activity kinetic profiles for both mutants with a synthetic small molecule substrate (pNPP) and the physiological relevant substrate (pERK) when compared to wild type GST-DUSP5 protein. CONCLUSION: Our molecular modeling and biochemical studies combined demonstrate that enzymatic activity of phosphatases can be manipulated by mutating specific conserved amino acid residues in the catalytic site (phosphatase domain). This strategy could facilitate generation of small molecules that will serve as agonists/antagonists of DUSP5 activity.

Ab Initio Study Predicts That Enigmatic Isonitrosyl Fluoride Should Be Stable at Low Temperatures yet Unnoticeable Due to Its Photoreactivity.

Isonitrosyl fluoride F-ON remains an undetected molecule despite multiple attempts to generate it and successful identification of other isonitrosyl halides (X-ONs) via phototransformations of corresponding X-NOs. We investigated this problem using ab initio methods and found no evidence of instability of F-ON at low temperatures of 8-10 K. Instead, experimental observation of F-ON is likely challenged by the (1) different nature of photoexcitation of F-NO and its quantum yield being lower than those of other X-NOs and (2) the presence of a bright charge-transfer transition in the F-ON spectrum that likely overlaps with the weak band of F-NO used for photoexcitation. Formation of F-ON via symmetry-prohibited photoexcitation of F-NO is followed by its immediate photodecomposition to the charge-transfer excited state and its conversion to F-NO upon de-excitation. Thus, F-ON should be readily observable using non-photochemistry methods such as microwave spectroscopy.