Structural determinants for activation of the Tau kinase CDK5 by the serotonin receptor 5-HT7R.

BACKGROUND: Multiple neurodegenerative diseases are induced by the formation and deposition of protein aggregates. In particular, the microtubule-associated protein Tau leads to the development of so-called tauopathies characterized by the aggregation of hyperphosphorylated Tau within neurons. We recently showed that the constitutive activity of the serotonin receptor 7 (5-HT7R) is required for Tau hyperphosphorylation and aggregation through activation of the cyclin-dependent kinase 5 (CDK5). We also demonstrated physical interaction between 5-HT7R and CDK5 at the plasma membrane suggesting that the 5-HT7R/CDK5 complex is an integral part of the signaling network involved in Tau-mediated pathology. METHODS: Using biochemical, microscopic, molecular biological, computational and AI-based approaches, we investigated structural requirements for the formation of 5-HT7R/CDK5 complex. RESULTS: We demonstrated that 5-HT7R domains responsible for coupling to Gs proteins are not involved in receptor interaction with CDK5. We also created a structural model of the 5-HT7R/CDK5 complex and refined the interaction interface. The model predicted two conserved phenylalanine residues, F278 and F281, within the third intracellular loop of 5-HT7R to be potentially important for complex formation. While site-directed mutagenesis of these residues did not influence Gs protein-mediated receptor signaling, replacement of both phenylalanines by alanine residues significantly reduced 5-HT7R/CDK5 interaction and receptor-mediated CDK5 activation, leading to reduced Tau hyperphosphorylation and aggregation. Molecular dynamics simulations of 5-HT7R/CDK5 complex for wild-type and receptor mutants confirmed binding interface stability of the initial model. CONCLUSIONS: Our results provide a structural basis for the development of novel drugs targeting the 5-HT7R/CDK5 interaction interface for the selective treatment of Tau-related disorders, including frontotemporal dementia and Alzheimer's disease.

Systematic discovery of protein interaction interfaces using AlphaFold and experimental validation.

Structural resolution of protein interactions enables mechanistic and functional studies as well as interpretation of disease variants. However, structural data is still missing for most protein interactions because we lack computational and experimental tools at scale. This is particularly true for interactions mediated by short linear motifs occurring in disordered regions of proteins. We find that AlphaFold-Multimer predicts with high sensitivity but limited specificity structures of domain-motif interactions when using small protein fragments as input. Sensitivity decreased substantially when using long protein fragments or full length proteins. We delineated a protein fragmentation strategy particularly suited for the prediction of domain-motif interfaces and applied it to interactions between human proteins associated with neurodevelopmental disorders. This enabled the prediction of highly confident and likely disease-related novel interfaces, which we further experimentally corroborated for FBXO23-STX1B, STX1B-VAMP2, ESRRG-PSMC5, PEX3-PEX19, PEX3-PEX16, and SNRPB-GIGYF1 providing novel molecular insights for diverse biological processes. Our work highlights exciting perspectives, but also reveals clear limitations and the need for future developments to maximize the power of Alphafold-Multimer for interface predictions.

High-Performance Hydrogel Sensors Enabled Multimodal and Accurate Human-Machine Interaction System for Active Rehabilitation.

Human-machine interaction (HMI) technology shows an important application prospect in rehabilitation medicine, but it is greatly limited by the unsatisfactory recognition accuracy and wearing comfort. Here, this work develops a fully flexible, conformable, and functionalized multimodal HMI interface consisting of hydrogel-based sensors and a self-designed flexible printed circuit board. Thanks to the component regulation and structural design of the hydrogel, both electromyogram (EMG) and forcemyography (FMG) signals can be collected accurately and stably, so that they are later decoded with the assistance of artificial intelligence (AI). Compared with traditional multichannel EMG signals, the multimodal human-machine interaction method based on the combination of EMG and FMG signals significantly improves the efficiency of human-machine interaction by increasing the information entropy of the interaction signals. The decoding accuracy of the interaction signals from only two channels for different gestures reaches 91.28%. The resulting AI-powered active rehabilitation system can control a pneumatic robotic glove to assist stroke patients in completing movements according to the recognized human motion intention. Moreover, this HMI interface is further generalized and applied to other remote sensing platforms, such as manipulators, intelligent cars, and drones, paving the way for the design of future intelligent robot systems.

Mapping Molecular Interaction Interface Between Diaphanous Formin-2 and Neuron-Specific Drebrin A.

Actin cytoskeleton is critical for neuronal shape and function. Drebrin and formins are key regulators of neuronal actin networks. Neuron-specific drebrin A is highly enriched in dendritic spines (postsynaptic terminals) of mature excitatory neurons. Decreased levels of drebrin in dendritic spines is a hallmark of Alzheimer's disease, epilepsy, and other complex disorders, which calls for better understanding of its regulatory functions. Drebrin A was previously shown to inhibit actin nucleation and bundling by the diaphanous formin-2 (mDia2) - an actin nucleator that is involved in the initiation of dendritic spines. Characterization of the molecular binding interface between mDia2 and drebrin is necessary to better understand the functional consequences of this interaction and its biological relevance. Prior work suggested a multi-pronged interface between mDia2 and drebrin, which involves both N-terminal and C-terminal regions of the drebrin molecule. Here we used mass spectrometry analysis, deletion mutagenesis, and an array of synthetic peptides of neuronal drebrin A to map its formin-binding interface. The mDia2-interacting interface on drebrin was narrowed down to three highly conserved 9-16 residue sequences that were used to identify some of the key residues involved in this interaction. Deletion of the C-terminal region of drebrin greatly reduces its binding to mDia2 and the extent of its inhibition of formin-driven actin assembly. Moreover, our experiments with formins from different subfamilies showed that drebrin is a specific rather than general inhibitor of these proteins. This work contributes to a molecular level understanding of the formin-drebrin interaction and will help to unravel its biological significance.

Enhancing Human-Robot Collaboration through a Multi-Module Interaction Framework with Sensor Fusion: Object Recognition, Verbal Communication, User of Interest Detection, Gesture and Gaze Recognition.

With the increasing presence of robots in our daily lives, it is crucial to design interaction interfaces that are natural, easy to use and meaningful for robotic tasks. This is important not only to enhance the user experience but also to increase the task reliability by providing supplementary information. Motivated by this, we propose a multi-modal framework consisting of multiple independent modules. These modules take advantage of multiple sensors (e.g., image, sound, depth) and can be used separately or in combination for effective human-robot collaborative interaction. We identified and implemented four key components of an effective human robot collaborative setting, which included determining object location and pose, extracting intricate information from verbal instructions, resolving user(s) of interest (UOI), and gesture recognition and gaze estimation to facilitate the natural and intuitive interactions. The system uses a feature-detector-descriptor approach for object recognition and a homography-based technique for planar pose estimation and a deep multi-task learning model to extract intricate task parameters from verbal communication. The user of interest (UOI) is detected by estimating the facing state and active speakers. The framework also includes gesture detection and gaze estimation modules, which are combined with a verbal instruction component to form structured commands for robotic entities. Experiments were conducted to assess the performance of these interaction interfaces, and the results demonstrated the effectiveness of the approach.

Serine pseudoproteases in physiology and disease.

Serine proteases (SPs) constitute a very important family of enzymes, both physiologically and pathologically. The effects produced by these proteins have been explained by their proteolytic activity. However, the discovery of pharmacologically active SP molecules that show no enzymatic activity, as the so-called pseudo SPs or SP homologs (SPHs), has exposed a profoundly neglected possibility of nonenzymatic functions of these SP molecules. In this review, the most thoroughly described SPHs are presented. The main physiological domains in which SPHs operate appear to be in reproduction, embryonic development, immune response, host defense, and hemostasis. Hitherto unexplained actions of SPs should therefore be considered also as the result of the ligand-like attributes of SPs. The gain of a novel function by an SPH is a consequence of specific amino acid replacements that have resulted in a novel interaction interface or a 'catalytic trap'. Unraveling the SP/SPH interactome will provide a description of previously unknown physiological functions of SPs/SPHs, aiding the creation of innovative medical approaches.

Impact of Asp/Glu-ADP-ribosylation on protein-protein interaction and protein function.

PARylation plays critical role in regulating multiple cellular processes such as DNA damage response and repair, transcription, RNA processing, and stress response. More than 300 human proteins have been found to be modified by PARylation on acidic residues, that is, Asp (D) and Glu (E). We used the deep-learning tool AlphaFold to predict protein-protein interactions (PPIs) and their interfaces for these proteins based on coevolution signals from joint multiple sequence alignments (MSAs). AlphaFold predicted 260 confident PPIs involving PARylated proteins, and about one quarter of these PPIs have D/E-PARylation sites in their predicted PPI interfaces. AlphaFold predictions offer novel insights into the mechanisms of PARylation regulations by providing structural details of the PPI interfaces. D/E-PARylation sites have a preference to occur in coil regions and disordered regions, and PPI interfaces containing D/E-PARylation sites tend to occur between short linear sequence motifs in disordered regions and globular domains. The hub protein PCNA is predicted to interact with more than 20 proteins via the common PIP box motif and the structurally variable flanking regions. D/E-PARylation sites were found in the interfaces of key components of the RNA transcription and export complex, the SF3a spliceosome complex, and H/ACA and C/D small nucleolar ribonucleoprotein complexes, suggesting that systematic PARylation have a profound effect in regulating multiple RNA-related processes such as RNA nuclear export, splicing, and modification. Finally, PARylation of SUMO2 could modulate its interaction with CHAF1A, thereby representing a potential mechanism for the cross-talk between PARylation and SUMOylation in regulation of chromatin remodeling.

Docking and Molecular Dynamics-Based Identification of Interaction between Various Beta-Amyloid Isoforms and RAGE Receptor.

Beta-amyloid peptide (Aß) is a ligand associated with RAGE (Advanced glycosylation end product-specific receptor). Aß is translocated in complexes with RAGE from the blood to brain across the blood-brain barrier (BBB) by transcytosis. Aß and its isoforms are important factors in the Alzheimer's disease (AD) pathogenesis. However, interaction with RAGE was previously studied for Aß but not for its isoforms. The present study has been directed at identifying the key interaction interfaces between RAGE and Aß isoforms (Aß40, Aß42, phosphorylated and isomerized isoforms pS8-Aß42, isoD7-Aß42). Two interfaces have been identified by docking: they are represented by an extended area at the junction of RAGE domains V and C1 and a smaller area linking C1 and C2 domains. Molecular dynamics (MD) simulations have shown that all Aß isoforms form stable and tightly bound complexes. This indicates that all Aß isoforms potentially can be transported through the cell as part of a complex with RAGE. Modeling of RAGE interaction interfaces with Aß indicates which chemical compounds can potentially be capable of blocking this interaction, and impair the associated pathogenic cascades. The ability of three RAGE inhibitors (RAP, FPS-ZM1 and RP-1) to disrupt the RAGE:Aß interaction has been probed by docking and subsequently the complexes' stability verified by MD. The RP-1 and Aß interaction areas coincide and therefore this inhibitor is very promising for the RAGE:Aß interaction inhibition.

Interaction Interface of Aß42 with Human Na,K-ATPase Studied by MD and ITC and Inhibitor Screening by MD.

Alzheimer's disease (AD) is a neurodegenerative disease accompanied by progressive cognitive and memory dysfunction due to disruption of normal electrotonic properties of neurons and neuronal loss. The Na,K-ATPase interaction with beta amyloid (Aß) plays an important role in AD pathogenesis. It has been shown that Na,K-ATPase activity in the AD brain was significantly lower than those in age-matched control brain. The interaction of Aß42 with Na,K-ATPase and subsequent oligomerization leads to inhibition of the enzyme activity. In this study interaction interfaces between three common Aß42 isoforms, and different conformations of human Na,K-ATPase (α1ß1) have been obtained using molecular modeling, including docking and molecular dynamics (MD). Interaction sites of Na,K-ATPase with Aß42 are localized between extracellular parts of α- and ß- subunits and are practically identical for Na,K-ATPase at different conformations. Thermodynamic parameters for the formation of Na,K-ATPase:Aß42 complex at different conformations acquired by isothermal titration calorimetry (ITC) are similar, which is in line with the data of molecular modeling. Similarity of Na,K-ATPase interaction interfaces with Aß in all conformations allowed us to cross-screen potential inhibitors for this interaction and find pharmaceutical compounds that could block it.

The Impact of Cognition on Motor Learning and Skill Acquisition Using a Robot Intervention in Infants With Cerebral Palsy.

Background: Cerebral Palsy (CP) is a neurodevelopmental disorder that encompasses multiple neurological disorders that appear in infancy or early childhood and persist through the lifespan of the individual. Early interventions for infants with CP utilizing assisted-motion robotic devices have shown promising effects in rehabilitation of the motor function skills. The impact of cognitive function during motor learning and skill acquisition in infants using robotic technologies is unclear. Purpose: To assess the impact of cognitive function of infants with and without CP on their motor learning using the Self-Initiated Prone Progression Crawler (SIPPC) robot. Methods: Statistical analysis was conducted on the data obtained from a randomized control trial in which the movement learning strategies in infants with or at risk for CP was assessed during a 16-week SIPPC robot intervention. Cognitive function was measured by the Bayley scales of Infant and Toddler Development-Third edition (Bayley-III) and motor function was measured by the Movement Observation Coding Scheme (MOCS). The infants were categorized into three distinct groups based on their cognitive scores at baseline: "above average" (n1 = 11), "below average" (n2 = 10), and "average" (n3 = 26). Tri-weekly averages of the MOCS scores (observations at five time points) were used for the analyses. This study involved computing descriptive statistics, data visualization, repeated measures analysis of variances (rmANOVA), and survival analyses. Results: The descriptive statistics were calculated for the MOCS and Bayley III scores. The repeated measures ANOVAs revealed that there was a statistically significant effect of time (p < 0.0001) on scores of all subscales of the MOCS. A statistically significant effect of interaction between group and time (p < 0.05) was found in MOCS scores of subscales 1 and 2. The survival analyses indicated that infants in different cognition groups significantly differed (p < 0.0001) in their ability to achieve the crawling milestone within the 16-week intervention period. Conclusion: The findings in this study reveal the key movement strategies required to move the SIPPC robot, assessed by the MOCS, vary depending on the infants' cognition. The SIPPC robot is well-matched to cognitive ability of infants with CP. However, lower cognitive ability was related to delayed improvement in their motor skills.

Biochemical and structural analyses reveal critical residues in δ subunit affecting its bindings to ß' subunit of Staphylococcus aureus RNA polymerase.

A large class of bacterial RNA polymerase (RNAP) from low-G + C-content Gram-positive bacterial strains, such as the major human pathogen Staphylococcus aureus, not only contain five conserved subunits (αI, αII, ß, ß' and ω), but also has a δ subunit. Despite being first identified as unique, Gram-positive specific component of RNAP apoenzyme more than 30 years ago and reported to be essential for transcription, the structural basis and molecular mechanism of δ subunit in the regulation of transcription remain poorly understood. Here, we performed structural analyses, site-directed mutagenesis and biochemical assays to uncover the interactions of S. aureus δ subunit with RNAP core enzyme and DNA towards the understanding of its role in transcription regulation. Microscale thermophoresis (MST) and electrophoretic mobility shift assay (EMSA) of the wild-type and mutated S. aureus δ subunit revealed the N-terminal domain of δ subunit directly binds to the ß' jaw of S. aureus RNAP (SauRNAP), identified the key amino acid residues (F58, D61, D65, R67 and W81) of δ subunit involving in the binding with SauRNAP core enzyme, and uncovered the δ subunit C-terminal domain interferes with the interaction between DNA and SauRNAP core enzyme, by which transcription is regulated. Our results provide an excellent starting point for understanding the unique regulatory role and physiological function of δ subunit on transcription regulation in Gram-positive bacteria.

Geometrical and electro-static determinants of protein-protein interactions.

Protein-protein interactions (PPI) are pivotal to the numerous processes in the cell. Therefore, it is of interest to document the analysis of these interactions in terms of binding sites, topology of the interacting structures and physiochemical properties of interacting interfaces and the of forces interactions. The interaction interface of obligatory protein-protein complexes differs from that of the transient interactions. We have created a large database of protein-protein interactions containing over100 thousand interfaces. The structural redundancy was eliminated to obtain a non-redundant database of over 2,265 interaction interfaces. Therefore, it is of interest to document the analysis of these interactions in terms of binding sites, topology of the interacting structures and physiochemical properties of interacting interfaces and the offorces interactions. The residue interaction propensity and all of the rest of the parametric scores converged to a statistical indistinguishable common sub-range and followed the similar distribution trends for all three classes of sequence-based classifications PPInS. This indicates that the principles of molecular recognition are dependent on the preciseness of the fit in the interaction interfaces. Thus, it reinforces the importance of geometrical and electrostatic complementarity as the main determinants for PPIs.

Assessment of Augmented Reality in Manual Wiring Production Process with Use of Mobile AR Glasses.

Digitalization of production environment, also called Industry 4.0 (the term invented by Wahlster Wolfgang in Germany) is now one of the hottest topics in the computer science departments at universities and companies. One of the most significant topics in this area is augmented reality (AR). The interest in AR has grown especially after the introduction of the Microsoft HoloLens in 2016, which made this technology available for researchers and developers all around the world. It is divided into numerous subtopics and technologies. These wireless, see-through glasses give a very natural human-machine interface, with the possibility to present certain necessary information right in front of the user's eyes as 3D virtual objects, in parallel with the observation of the real world, and the possibility to communicate with the system by simple gestures and speech. Scientists noted that in-depth studies connected to the effects of AR applications are presently sparse. In the first part of this paper, the authors recall the research from 2019 about the new method of manual wiring support with the AR glasses. In the second part, the study (tests) for this method carried out by the research team is described. The method was applied in the actual production environment with consideration of the actual production process, which is manual wiring of the industrial enclosures (control cabinets). Finally, authors deliberate on conclusions, technology's imperfections, limitations, and future possible development of the presented solution.

Structural Mechanism of the Arrestin-3/JNK3 Interaction.

Arrestins, in addition to desensitizing GPCR-induced G protein activation, also mediate G protein-independent signaling by interacting with various signaling proteins. Among these, arrestins regulate MAPK signal transduction by scaffolding mitogen-activated protein kinase (MAPK) signaling components such as MAPKKK, MAPKK, and MAPK. In this study, we investigated the binding mode and interfaces between arrestin-3 and JNK3 using hydrogen/deuterium exchange mass spectrometry, 19F-NMR, and tryptophan-induced Atto 655 fluorescence-quenching techniques. Results suggested that the ß1 strand of arrestin-3 is the major and potentially only interaction site with JNK3. The results also suggested that C-lobe regions near the activation loop of JNK3 form the potential binding interface, which is variable depending on the ATP binding status. Because the ß1 strand of arrestin-3 is buried by the C-terminal strand in its basal state, C-terminal truncation (i.e., pre-activation) of arrestin-3 facilitates the arrestin-3/JNK3 interaction.

Modeling of Protein⁻Protein Interactions in Cytokinin Signal Transduction.

The signaling of cytokinins (CKs), classical plant hormones, is based on the interaction of proteins that constitute the multistep phosphorelay system (MSP): catalytic receptors-sensor histidine kinases (HKs), phosphotransmitters (HPts), and transcription factors-response regulators (RRs). Any CK receptor was shown to interact in vivo with any of the studied HPts and vice versa. In addition, both of these proteins tend to form a homodimer or a heterodimeric complex with protein-paralog. Our study was aimed at explaining by molecular modeling the observed features of in planta protein-protein interactions, accompanying CK signaling. For this purpose, models of CK-signaling proteins' structure from Arabidopsis and potato were built. The modeled interaction interfaces were formed by rather conserved areas of protein surfaces, complementary in hydrophobicity and electrostatic potential. Hot spots amino acids, determining specificity and strength of the interaction, were identified. Virtual phosphorylation of conserved Asp or His residues affected this complementation, increasing (Asp-P in HK) or decreasing (His-P in HPt) the affinity of interacting proteins. The HK-HPt and HPt-HPt interfaces overlapped, sharing some of the hot spots. MSP proteins from Arabidopsis and potato exhibited similar properties. The structural features of the modeled protein complexes were consistent with the experimental data.

Characterization of the interaction interface and conformational dynamics of human TGIF1 homeodomain upon the binding of consensus DNA.

The TG interacting factor-1 homeodomain (TGIF1-HD) binds with the consensus DNA motif 5'-TGTCA-3' in gene promoters through its three-amino acid loop extension (TALE) type homeodomain, and then recruits co-regulators to regulate gene expression. Although the solution NMR structure of human TGIF1-HD has been reported previously, little is known about its DNA binding mechanism. NMR titrations have been extensively used to study mechanisms of ligand binding to target proteins; however, an intermediate exchange occurred predominantly between TGIF1-HD in the free and bound states when titrated with the consensus DNA, which resulted in poor-quality NMR spectra and precluded further exploration of its interaction interface and conformational dynamics. Here, the helix α3 of TGIF1-HD was speculated as the specific DNA binding interface by hydrogen-deuterium exchange mass spectrometry (HDX-MS) experiments, and subsequently confirmed by chemical exchange saturation transfer (CEST) spectroscopy. In addition, simultaneous conformational changes in other regions, including α1 and α2, were induced by DNA binding, explaining the observation of chemical shift perturbations from extensive residues besides those located in α3. Further, low-populated DNA-bound TGIF1-HD undergoing a slow exchange at a rate of 130.2 ±â€¯3.6 s-1 was derived from the analysis of the CEST data, and two residues, R220 and R221, located in the middle of α3 were identified to be crucial for DNA binding. Our study provides structural and dynamic insights into the mechanisms of TGIF1-HD recognition of extensive promoter DNA.

Modeling of RAS complexes supports roles in cancer for less studied partners.

BACKGROUND: RAS protein interactions have predominantly been studied in the context of the RAF and PI3kinase oncogenic pathways. Structural modeling and X-ray crystallography have demonstrated that RAS isoforms bind to canonical downstream effector proteins in these pathways using the highly conserved switch I and II regions. Other non-canonical RAS protein interactions have been experimentally identified, however it is not clear whether these proteins also interact with RAS via the switch regions. RESULTS: To address this question we constructed a RAS isoform-specific protein-protein interaction network and predicted 3D complexes involving RAS isoforms and interaction partners to identify the most probable interaction interfaces. The resulting models correctly captured the binding interfaces for well-studied effectors, and additionally implicated residues in the allosteric and hyper-variable regions of RAS proteins as the predominant binding site for non-canonical effectors. Several partners binding to this new interface (SRC, LGALS1, RABGEF1, CALM and RARRES3) have been implicated as important regulators of oncogenic RAS signaling. We further used these models to investigate competitive binding and multi-protein complexes compatible with RAS surface occupancy and the putative effects of somatic mutations on RAS protein interactions. CONCLUSIONS: We discuss our findings in the context of RAS localization to the plasma membrane versus within the cytoplasm and provide a list of RAS protein interactions with possible cancer-related consequences, which could help guide future therapeutic strategies to target RAS proteins.

Mapping Region of Human Restriction Factor APOBEC3H Critical for Interaction with HIV-1 Vif.

The APOBEC3 (A3) family of cellular cytidine deaminases comprises seven members (A, B, C, D, F, G, and H) that potently inhibit retroviral replication. Human immunodeficiency virus type 1 (HIV-1) Vif is a small pleiotropic protein that specifically inactivates these enzymes, targeting them for ubiquitin-mediated proteasomal degradation. A3 Vif-interaction sites are presumed to fall into three distinct types: A3C/D/F, A3G, and A3H. To date, two types of A3G and A3C/D/F sites have been well characterized, whereas the A3H Vif-binding site remains poorly defined. Here, we explore the residues critical for the A3H-type Vif interaction. To avoid technical difficulties in performing experiments with human A3H haplotype II (hapII), which is relatively resistant to HIV-1 Vif, we employed its ortholog chimpanzee A3H (cA3H), which displays high Vif sensitivity, for a comparison of sensitivity with that of A3H hapII. The Vif susceptibility of A3H hapII-cA3H chimeras and their substitution mutants revealed a single residue at position 97 as a major determinant for the difference in their Vif sensitivities. We further surveyed critical residues by structure-guided mutagenesis using an A3H structural model and thus identified eight additional residues important for Vif sensitivity, which mapped to the α3 and α4 helices of A3H. Interestingly, this area is located on a surface adjacent to the A3G and A3C/D/F interfaces and is composed of negatively charged and hydrophobic patches. These findings suggest that HIV-1 Vif has evolved to utilize three dispersed surfaces for recognizing three types of interfaces on A3 proteins under certain structural constraints.

Genetically encoded photocrosslinkers for identifying and mapping protein-protein interactions in living cells.

Protein-protein interactions (PPIs) play pivotal roles in regulation of many biological processes. Conventional methods are capable of investigating stable and strong interactions within protein complexes, but remain difficult for studying dynamic, transient, and weak PPIs. Herein we review photo-affinity unnatural amino acids that can be site-specifically incorporated into a protein of interest to covalently trap noncovalent PPIs under living conditions. A newly developed cleavable photocrosslinker from our group will also be introduced, which facilitated the prey-bait separation for better enrichment and identification of photocrosslinked PPI complexes. © 2016 IUBMB Life, 68(11):879-886, 2016.

Interaction between bound water molecules and local protein structures: A statistical analysis of the hydrogen bond structures around bound water molecules.

Water molecules play an important role in protein folding and protein interactions through their structural association with proteins. Examples of such structural association can be found in protein crystal structures, and can often explain protein functionality in the context of structure. We herein report the systematic analysis of the local structures of proteins interacting with water molecules, and the characterization of their geometric features. We first examined the interaction of water molecules with a large local interaction environment by comparing the preference of water molecules in three regions, namely, the protein-protein interaction (PPI) interfaces, the crystal contact (CC) interfaces, and the non-interfacial regions. High preference of water molecules to the PPI and CC interfaces was found. In addition, the bound water on the PPI interface was more favorably associated with the complex interaction structure, implying that such water-mediated structures may participate in the shaping of the PPI interface. The pairwise water-mediated interaction was then investigated, and the water-mediated residue-residue interaction potential was derived. Subsequently, the types of polar atoms surrounding the water molecules were analyzed, and the preference of the hydrogen bond acceptor was observed. Furthermore, the geometries of the structures interacting with water were analyzed, and it was found that the major structure on the protein surface exhibited planar geometry rather than tetrahedral geometry. Several previously undiscovered characteristics of water-protein interactions were unfolded in this study, and are expected to lead to a better understanding of protein structure and function.