Domain Interface Research Articles

Can Non-steroidal Anti-inflammatory Drugs Affect the Interaction Between Receptor Binding Domain of SARS-COV-2 Spike and the Human ACE2 Receptor? A Computational Biophysical Study

SARS-CoV-2 has caused millions of infections and more than 600,000 deaths worldwide. Despite the wide number of studies to date, there is no specifically effective treatment available for SARS-CoV-2. However, it has been proposed to target reused drugs with potential antiviral activity to the interfere between the angiotensin-converting enzyme 2 (ACE2) and the receptor binding domain (RBD) interface of SARS-CoV-2 to avoid cell recognition. Several non-steroidal anti-inflammatory drugs (NSAIDs) have been reported to have some type of activity against a wide variety of viruses including SARS-CoV-2. Therefore, we carried out an exhaustive computational biophysical study of various NSAIDs targeting the RBD-ACE2 complex using multiple comparative analysis of docking and molecular dynamics. Only the Ibuprofen (Propionic acid derivative), Aspirin (Salicylate), and the Acetaminophen (p-aminophenol derivative) had a thermodynamically favorable docking with the complex RBD-ACE2 interface under the conditions of this study. Although, Ibuprofen was the NSAIDs most thermodynamically favorable docking in the shortest simulation time, and was the major inducer of structural changes, conformational changes, and overall changes in the complex throughout the simulation, including disturbances in composition and distribution of cavities at the interface. Results that point to ibuprofen as an NSAID that, under the conditions outlined in this investigation, may have the highest probability of generating a disturbance in the stability of the RBD-ACE2 complex. This statement, although it could contribute information for the empirical treatment and prevention of COVID-19, represents only a theoretical orientation and approach, and requires its experimental demonstration because our predictions cannot secure a pharmacologically and clinically relevant interaction. Therefore, our results suggest a possible alternative mechanism of action of some NSAIDs against COVID-19 which is relevant because there are no reports related to this, in addition to their well-known anti-inflammatory properties.

Multi-scale three-domain approach for coupling free flow and flow in porous media including droplet-related interface processes

Drops on a free-flow/porous-medium-flow interface have a strong influence on the exchange of mass, momentum and energy between the two macroscopic flow regimes. Modeling droplet-related pore-scale processes in a macro-scale context is challenging due to the scale gap, but might be rewarding due to relatively low computational costs. We develop a three-domain approach to model drop formation, growth, detachment and film flow in a lower-dimensional interface domain. A simple upscaling technique allows to compute the drop-covered interface area fraction which affects the coupling fluxes. In a first scenario, only drop formation, growth and detachment are taken into account. Then, spreading and merging due to lateral fluxes are considered as well. The simulation results show that the impact of these droplet-related processes can be captured. However, extensions are necessary to represent the influence on the free flow more precisely.

Uncaria tomentosa (cat’s claw): a promising herbal medicine against SARS-CoV-2/ACE-2 junction and SARS-CoV-2 spike protein based on molecular modeling

COVID-19 is a novel severe acute respiratory syndrome coronavirus. Currently, there is no effective treatment and vaccines seem to be the solution in the future. Virtual screening of potential drugs against the S protein of severe acute respiratory syndrome corona virus 2 (SARS-CoV-2) has provided small molecular compounds with a high binding affinity. Unfortunately, most of these drugs do not attach with the binding interface of the receptor-binding domain (RBD)–angiotensin-converting enzyme-2 (ACE-2) complex in host cells. Molecular modeling was carried out to evaluate the potential antiviral properties of the components of the medicinal herb Uncaria tomentosa (cat’s claw) focusing on the binding interface of the RBD–ACE-2 and the viral spike protein. The in silico approach starts with protein–ligand docking of 26 Cat’s claw key components followed by molecular dynamics simulations and re-docked calculations. Finally, we carried out drug-likeness calculations for the most qualified cat’s claw components. The structural bioinformatics approaches led to the identification of several bioactive compounds of U. tomentosa with potential therapeutic effect by dual strong interaction with interface of the RBD–ACE-2 and the ACE-2 binding site on SARS-CoV-2 RBD viral spike. In addition, in silico drug-likeness indices for these components were calculated and showed good predicted therapeutic profiles of these phytochemicals found in U. tomentosa (cat’s claw). Our findings suggest the potential effectiveness of cat’s claw as complementary and/or alternative medicine for COVID-19 treatment. Communicated by Ramaswamy H. Sarma

Autophosphorylation of EGFR at Y954 Facilitated Homodimerization and Enhanced Downstream Signals

Asymmetric dimer formation of epidermal growth factor receptor (EGFR) is crucial for EGF-induced receptor activation. Even though autophosphorylation is important for activation, its role remains elusive in the context of regulating dimers. In this study, employing overlapping time series analysis to raster image correlation spectroscopy (RICS), we observed time-dependent transient dynamics of EGFR dimerization and found EGFR kinase activity to be essential for dimerization. As a result of which, we hypothesized that phosphorylation could influence dimerization. Evaluating this point, we observed that one of the tyrosine residues (Y954) located in the C-terminal lobe of the activator kinase domain was important to potentiate dimerization. Functional imaging to monitor Ca2+ and ERK signals revealed a significant role of Y954 in influencing downstream signaling cascade. Crucial for stabilization of EGFR asymmetric dimer is a “latch” formed between kinase domains of the binding partners. Because Y954 is positioned adjacent to the latch binding region on the kinase domain, we propose that phosphorylation strengthened the latch interaction. On the contrary, we identified that threonine phosphorylation (T669) in the latch domain negatively regulated EGFR dimerization and the downstream signals. Overall, we have delineated the previously anonymous role of phosphorylation at the latch interface of kinase domains in regulating EGFR dimerization.

The NAD+-mediated self-inhibition mechanism of pro-neurodegenerative SARM1.

Pathological degeneration of axons disrupts neural circuits and represents one of the hallmarks of neurodegeneration1-4. Sterile alpha and Toll/interleukin-1 receptor motif-containing protein 1 (SARM1) is a central regulator of this neurodegenerative process5-8, and its Toll/interleukin-1 receptor (TIR) domain exerts its pro-neurodegenerative action through NADase activity9,10. However, the mechanisms by which the activation of SARM1 is stringently controlled are unclear. Here we report the cryo-electron microscopy structures of full-length SARM1 proteins. We show that NAD+ is an unexpected ligand of the armadillo/heat repeat motifs (ARM) domain of SARM1. This binding of NAD+ to the ARM domain facilitated the inhibition of the TIR-domain NADase through the domain interface. Disruption of the NAD+-binding site or the ARM-TIR interaction caused constitutive activation of SARM1 and thereby led to axonal degeneration. These findings suggest that NAD+ mediates self-inhibition of this central pro-neurodegenerative protein.

Molecular dynamics simulation study on Thermotoga maritima EngA: GTP/GDP bound state of the second G-domain influences the domain–domain interface interactions

EngA, a GTPase involved in the late steps of ribosome maturation, consists of two GTP binding domains (G-domains) [GD1, GD2] and a C-terminal domain. The combination of GTP/GDP in G-domains dictates its binding to the ribosomal subunits by altering its conformation. Studies and comparisons on the available structures of EngA enable us to understand the correlation between nucleotide bound states and its conformation. Using all-atom molecular dynamics (MD) simulations, we have explored the conformational behavior of EngA from Thermotoga maritima (TmDer) upon binding the various combinations of GTP and GDP. Analyses of Root Mean Square Deviation (RMSD), Radius of Gyration (Rg) and Root Mean Square Fluctuation (RMSF) emphasize the importance of the second G-domain nucleotide bound state. RMSD and Rg exhibit slightly lower values when GTP is embedded in GD2 compared to GDP. These lower values are due to Sw-II of GD2, which has been observed from RMSF plot. Further investigation on the effects of GD2 nucleotide bound state using Principal Component Analysis (PCA) and Free Energy Landscape (FEL) analysis manifests an allosteric connection between GD2 nucleotide bound state and the GD1–KH interface. This is further validated by extracting electrostatic interactions and H-bonds at the GD1–KH interface. In silico mutations at the GD1 interface of KH domain affect the Sw-II mobility of GD2 by showing inverted behavior. This suggests using the second G-domain as an antibacterial target and further simulation studies on different species of EngA are to be explored. Communicated by Ramaswamy H. Sarma

Solution structure of the Hop TPR2A domain and investigation of target druggability by NMR, biochemical and in silico approaches

Heat shock protein 90 (Hsp90) is a molecular chaperone that plays an important role in tumour biology by promoting the stabilisation and activity of oncogenic ‘client’ proteins. Inhibition of Hsp90 by small-molecule drugs, acting via its ATP hydrolysis site, has shown promise as a molecularly targeted cancer therapy. Owing to the importance of Hop and other tetratricopeptide repeat (TPR)-containing cochaperones in regulating Hsp90 activity, the Hsp90-TPR domain interface is an alternative site for inhibitors, which could result in effects distinct from ATP site binders. The TPR binding site of Hsp90 cochaperones includes a shallow, positively charged groove that poses a significant challenge for druggability. Herein, we report the apo, solution-state structure of Hop TPR2A which enables this target for NMR-based screening approaches. We have designed prototype TPR ligands that mimic key native ‘carboxylate clamp’ interactions between Hsp90 and its TPR cochaperones and show that they block binding between Hop TPR2A and the Hsp90 C-terminal MEEVD peptide. We confirm direct TPR-binding of these ligands by mapping 1H–15N HSQC chemical shift perturbations to our new NMR structure. Our work provides a novel structure, a thorough assessment of druggability and robust screening approaches that may offer a potential route, albeit difficult, to address the chemically challenging nature of the Hop TPR2A target, with relevance to other TPR domain interactors.

Nitrosylation of the Diiron Core Mediated by the N Domain of YtfE.

The YtfE protein catalyzes the reduction of NO to N2O, protecting iron-sulfur clusters from nitrosylation. The structure of YtfE has a two-domain architecture, with a diiron-containing C-terminal domain linked to an N-terminal domain, in which the function of the latter is enigmatic. Here, by using electron spin resonance (ESR) spectroscopy, we show that YtfE exists in two conformational states, one of which has not been reported. Under high osmotic stress, YtfE adopts a homogeneous conformation (C state) similar to the known crystal structure. In a regular buffer, the N-terminal domain switches between the C state and a previously unidentified conformation (C' state), the latter of which has more space at the domain interface to allow the trafficking of NO molecules and thus is proposed to be a functionally active state. The conformational switch between the C and C' states is pivotal for facilitating NO access to the diiron core.

A Conformational Escape Reaction of HIV-1 against an Allosteric Integrase Inhibitor.

HIV-1 often acquires drug-resistant mutations in spite of the benefits of antiretroviral therapy (ART). HIV-1 integrase (IN) is essential for the concerted integration of HIV-1 DNA into the host genome. IN further contributes to HIV-1 RNA binding, which is required for HIV-1 maturation. Non-catalytic-site integrase inhibitors (NCINIs) have been developed as allosteric IN inhibitors, which perform anti-HIV-1 activity by a multimodal mode of action such as inhibition of the IN-lens epithelium-derived growth factor (LEDGF)/p75 interaction in the early stage and disruption of functional IN multimerization in the late stage of HIV-1 replication. Here, we show that IN undergoes an adaptable conformational change to escape from NCINIs. We observed that NCINI-resistant HIV-1 variants have accumulated 4 amino acid mutations by passage 26 (P26) in the IN-encoding region. We employed high-performance liquid chromatography (HPLC), thermal stability assays, and X-ray crystallographic analysis to show that some amino acid mutations affect the stability and/or dimerization interface of the IN catalytic core domains (CCDs), potentially resulting in the severely decreased multimerization of full-length IN proteins (IN undermultimerization). This undermultimerized IN via NCINI-related mutations was stabilized by HIV-1 RNA and restored to the same level as that of wild-type HIV-1 in viral particles. Recombinant HIV-1 clones with IN undermultimerization propagated similarly to wild-type HIV-1. Our study revealed that HIV-1 can eventually counteract NCINI-induced IN overmultimerization by IN undermultimerization as one of the escape mechanisms. Our findings provide information on the understanding of IN multimerization with or without HIV-1 RNA and may influence the development of anti-HIV-1 strategies.IMPORTANCE Understanding the mechanism of HIV-1 resistance to anti-HIV-1 drugs could lead to the development of novel drugs with increased efficiency, resulting in more effective ART. ART composed of more potent and long-acting anti-HIV-1 drugs can greatly improve drug adherence and also provide HIV-1 prevention such as preexposure prophylaxis. NCINIs with a multimodal mode of action exert potent anti-HIV-1 effects through IN overmultimerization during HIV-1 maturation. However, HIV-1 can acquire some mutations that cause IN undermultimerization to alleviate NCINI-induced IN overmultimerization. This undermultimerized IN was efficiently stabilized by HIV-1 RNA and restored to the same level as that of wild-type HIV-1. Our findings revealed that HIV-1 eventually acquires such a conformational escape reaction to overcome the unique NCINI actions. The investigation into drug-resistant mutations associated with HIV-1 protein multimerization may facilitate the elucidation of its molecular mechanism and functional multimerization, allowing us to develop more potent anti-HIV-1 drugs and unique treatment strategies.

Interface magnetoelectric effect and its sensitivity on interface structures in Fe/AgNbO3 and SrRuO3/AgNbO3 heterostructures: A first-principles investigation

The interface magnetoelectric (ME) effect is promising for novel technological applications as it allows the control of interface magnetization at heterointerfaces by the external electric field. Here in this article, we explore the interface ME effect and its sensitivity on interface atomic structure and bonding in ferromagnetic-ferroelectric oxide hetrostructures within the framework of density functional theory. In particular, we consider Fe/AgNbO3 and SrRuO3/AgNbO3 superlattices with different possible defect-free interfaces. Our results suggest that interface magnetization and thereby interface ME effect can critically depend on interface structure. The interface magnetization is found to depend on atomic bondings which are sensitive to atomic displacements at the interface. This leads to ME coupling due to induced change in the interface magnetization as the electric polarization is reversed in the ferroelectric film. In addition, the contribution to interface ME coupling also comes from the change in exchange-splitting between spin-polarized electrons and consequently change in the interface magnetic moments due to ferroelectric polarization reversal. The change in exchange splitting, in turn, is caused by spin-dependent screening of the bound polarization charges at the interface. In Fe/AgNbO3 system, the interface ME coefficient is found to exhibit opposing trend for Fe|AgO and Fe|NbO2 interfaces. In addition to interface ME effect, the formation of ferroelectric interface domain wall (IDW) in these systems is also explored. The IDW is found to form only in SrRuO3/AgNbO3 system with SrO|NbO2 interface due to oppositely oriented strong dipole moments at the two interfaces. The strong dependence of interface ME effect on interface atomic structure may have implication for its experimental observation in oxide heterostructures. In particular, the interface defects may render the conclusive observation of this phenomenon challenging due to resulting fluctuations in the observed data.

Biochemical Characterization of Respiratory Syncytial Virus RNA-Dependent RNA Polymerase Complex.

RNA-dependent RNA polymerases (RdRPs) from nonsegmented negative strand (NNS) RNA viruses perform both mRNA transcription and genome replication, and these activities are regulated by their interactions with RNA and other accessory proteins within the ribonucleoprotein (RNP) complex. Detailed biochemical characterization of these enzymatic activities and their regulation is essential for understanding the life cycles of many pathogenic RNA viruses and for antiviral drug discovery. We developed biochemical and biophysical kinetic methods to study the RNA synthesis and RNA binding activities of respiratory syncytial virus (RSV) L/P RdRP. We determined that the intact L protein is essential for RdRP activity, and in truncated L protein constructs, RdRP activity is abrogated due to their deficiency in RNA template binding. These results are in agreement with the observation of an RNA template-binding tunnel at the interface of RdRP and capping domains in RSV and vesicular stomatitis virus (VSV) L protein cryo-EM structures. We also describe nonradiometric assays for measuring RNA binding and RNA polymerization activity of RSV RdRP, which are amenable to compound screening and profiling.

Method of Boundary Integral Equations in the Problem of Diffraction of a Monochromatic Electromagnetic Wave by a System of Perfectly Conducting and Piecewise Homogeneous Dielectric Objects

We consider the three-dimensional problem of diffraction of a monochromatic electromagnetic wave by a system of objects of various physical nature, including dielectric bodies (domains), perfectly conducting bodies, and perfectly conducting screens. The perfectly conducting bodies may be placed in an exterior medium or immersed into the dielectric domains. In addition, the perfectly conducting screens may reside at the interface of the dielectric domains, being part of each of those. We give a boundary value problem for the Maxwell equations that describes the electromagnetic field under consideration. Integral representations are derived for the electromagnetic field in terms of surface integrals, and the boundary value problem is reduced to a system of boundary integral equations containing weakly and strongly singular surface integrals. The integral equations are written on the dielectric and perfectly conducting parts of the interface between the dielectric domains, on the surfaces of the perfectly conducting bodies, and on the perfectly conducting screens.

Function-related asymmetry of the specific cardiolipin binding sites on the mitochondrial ADP/ATP carrier

The ADP/ATP carrier (AAC) transports matrix ATP and cytosolic ADP across the inner mitochondrial membrane (IMM). It is well known that cardiolipin (CL) plays an important role in regulating the function of AAC, yet the underlying mechanism still remains elusive. AAC is composed of three homologous domains, and three specific CL binding sites are located at the domain-domain interfaces near the matrix side. Here we report an in-depth investigation on the dynamic properties of the bound CL within the three specific sites through all-atom molecular dynamics simulations of up to 13 μs in total. Our results highlight the importance of the basic and polar residues in CL binding. The basic residues from the linker helix and/or the [Y/W/F][K/R]G motif enable the bound CL to form an intra-domain binding mode, and the canonical inter-domain binding mode only forms when these basic residues are occupied by an additional phospholipid. Of special significance, differences in the basic and polar residues lead to remarkable asymmetry among the three specific CL binding sites. We found that the bound CL at the interface of domains 2 and 3 predominantly adopts inter-domain binding mode, while CLs at the other two sites have much more intra-domain populations. This is consistent with the asymmetric crystal structure of the matrix state (m-state) AAC which implies an asymmetric transport mechanism. The dynamic equilibrium between the inter-domain and intra-domain binding modes observed in our simulations could be highly important for the bound CLs to adapt to the movements during state transitions.

Impaired cytoplasmic domain interactions cause co-assembly defect and loss of function in the p.Glu293Lys KNCJ2 variant isolated from an Andersen-Tawil syndrome patient.

Subunit interactions at the cytoplasmic domain interface (CD-I) have recently been shown to control gating in inward rectifier potassium channels. Here we report the novel KCNJ2 variant p.Glu293Lys that has been found in a patient with Andersen-Tawil syndrome type 1 (ATS1), causing amino acid substitution at the CD-I of the inward rectifier potassium channel subunit Kir2.1. Neither has the role of Glu293 in gating control been investigated nor has a pathogenic variant been described at this position. This study aimed to assess the involvement of Glu293 in CD-I subunit interactions and to establish the pathogenic role of the p.Glu293Lys variant in ATS1. The p.Glu293Lys variant produced no current in homomeric form and showed dominant-negative effect over wild-type (WT) subunits. Immunocytochemical labelling showed the p.Glu293Lys subunits to distribute in the subsarcolemmal space. Salt bridge prediction indicated the presence of an intersubunit salt bridge network at the CD-I of Kir2.1, with the involvement of Glu293. Subunit interactions were studied by the NanoLuc® Binary Technology (NanoBiT) split reporter assay. Reporter constructs carrying NanoBiT tags on the intracellular termini produced no bioluminescent signal above background with the p.Glu293Lys variant in homomeric configuration and significantly reduced signals in cells co-expressing WT and p.Glu293Lys subunits simultaneously. Extracellularly presented reporter tags, however, generated comparable bioluminescent signals with heteromeric WT and p.Glu293Lys subunits and with homomeric WT channels. Loss of function and dominant-negative effect confirm the causative role of p.Glu293Lys in ATS1. Co-assembly of Kir2.1 subunits is impaired in homomeric channels consisting of p.Glu293Lys subunits and is partially rescued in heteromeric complexes of WT and p.Glu293Lys Kir2.1 variants. These data point to an important role of Glu293 in mediating subunit assembly, as well as in gating of Kir2.1 channels.

A cryptic tubulin-binding domain links MEKK1 to curved tubulin protomers

The MEKK1 protein is a pivotal kinase activator of responses to cellular stress. Activation of MEKK1 can trigger various responses, including mitogen-activated protein (MAP) kinases, NF-κB signaling, or cell migration. Notably, MEKK1 activity is triggered by microtubule-targeting chemotherapies, among other stressors. Here we show that MEKK1 contains a previously unidentified tumor overexpressed gene (TOG) domain. The MEKK1 TOG domain binds to tubulin heterodimers-a canonical function of TOG domains-but is unusual in that it appears alone rather than as part of a multi-TOG array, and has structural features distinct from previously characterized TOG domains. MEKK1 TOG demonstrates a clear preference for binding curved tubulin heterodimers, which exist in soluble tubulin and at sites of microtubule polymerization and depolymerization. Mutations disrupting tubulin binding decrease microtubule density at the leading edge of polarized cells, suggesting that tubulin binding may play a role in MEKK1 activity at the cellular periphery. We also show that MEKK1 mutations at the tubulin-binding interface of the TOG domain recur in patient-derived tumor sequences, suggesting selective enrichment of tumor cells with disrupted MEKK1-microtubule association. Together, these findings provide a direct link between the MEKK1 protein and tubulin, which is likely to be relevant to cancer cell migration and response to microtubule-modulating therapies.

The Art of Swinging: How Boundary-Spanning Units Change Boundary Positions

In this study, we explore how boundary-spanning units positioned at the interface of different social domains change boundary positions over time in response to changing stakeholder demands. To do so, we draw on a longitudinal, inductive single case study of AppCampus, a boundary organization mediating collaboration between the corporate, university, and startup worlds. In examining how an initial boundary position evolves beyond the founding phase, we focus particularly on the actions and interactions of the organizational members of the boundary-spanning unit and its stakeholders. Our findings show how the boundary position evolves as a consequence of translation and acculturation. By illuminating how a boundary-spanning unit changes over time, we contribute to the literatures on boundary spanning and institutional theory.

Crystal structure of the FYCO1 RUN domain suggests possible interfaces with small GTPases.

FYCO1 is a multidomain adaptor protein that plays an important role in autophagy by mediating the kinesin-dependent microtubule plus-end-directed transport of autophagosomes. FYCO1 contains a RUN domain, which is hypothesized to function as a specific effector for members of the Ras superfamily of small GTPases, but its role has not been well characterized and its interaction partner(s) have not been identified. Here, the crystal structure of the FYCO1 RUN domain was determined at 1.3 Å resolution. The overall structure of the FYCO1 RUN domain was similar to those of previously reported RUN domains. Detailed structural comparisons with other RUN domains and docking studies suggested a possible interaction interface of the FYCO1 RUN domain with small GTPases of the Ras superfamily.

The Effects of Framework Mutations at the Variable Domain Interface on Antibody Affinity Maturation in an HIV-1 Broadly Neutralizing Antibody Lineage.

Understanding affinity maturation of antibodies that can target many variants of HIV-1 is important for vaccine development. While the antigen-binding site of antibodies is known to mutate throughout the co-evolution of antibodies and viruses in infected individuals, the roles of the mutations in the antibody framework region are not well understood. Throughout affinity maturation, the CH103 broadly neutralizing antibody lineage, from an individual designated CH505, altered the orientation of one of its antibody variable domains. The change in orientation was a response to insertions in the variable loop 5 (V5) of the HIV envelope. In this study, we generated CH103 lineage antibody variants in which residues in the variable domain interface were mutated, and measured the binding to both autologous and heterologous HIV-1 envelopes. Our data show that very few mutations in an early intermediate antibody of the lineage can improve binding toward both autologous and heterologous HIV-1 envelopes. We also crystallized an antibody mutant to show that framework mutations alone can result in a shift in relative orientations of the variable domains. Taken together, our results demonstrate the functional importance of residues located outside the antigen-binding site in affinity maturation.

Properties of cellulose reinforced composites: A review

This paper provides a review of cellulose, sources, extraction, molecular structure, cellulose whiskers, preparations, and morphology. The mechanical and thermal properties of cellulose reinforced composites are also discussed. Detail structure of Nano whiskers is also reported. As a renewable biomaterial, the most common source of cellulose is the plant. These plants include fruit fibers (coir), seed fibers (cotton), wood, leaf fibers (sisal), bast fibers (jute, kenaf, and hemp). Other sources of cellulose are from micro-organisms such as fungi, tunicates, bacteria, and algae. Cellulose whiskers are isolated from cellulose fibers by acid hydrolysis. Cellulose micro fibril structures are made of both amorphous and crystalline regions. The amorphous regions are vulnerable to hydrolysis by acids compared to the crystalline domains. Several techniques among which are Field Emission Scanning Electron Microscopy (FESEM), Transmission electron microscopy (TEM) and Atomic Force Microscopy (AFM) have been used to study the morphology of cellulose whiskers. An interface between cellulose whisker and matrix is a transition zone between the matrix and the cellulose whiskers. It plays an important role in the overall mechanical properties of the composites. A soft interface domain will yield a greater resistance to fracture, while the composite will be low in stiffness and strength. On the other hand, a stiffer interface domain may cause the composite to be strong and stiff and less resistant to fracture. The addition of CW into polymers matrices has little or no effect on the glass transition temperature, (Tg) except on the modification of CW.
 Keywords: Cellulose, whiskers, mechanical, thermal, properties, biomaterials

Rationally engineered prolyl endopeptidases from Sphingomonas capsulata with improved hydrolytic activity towards pathogenic peptides of celiac diseases

Celiac disease affects approximately 1% of the population and is a major public health problem worldwide. It is trigged by gluten-derived peptides, which have unusually high proline-glutamine motif content and are highly resistant to proteolysis by digestive enzymes of the gastrointestinal tract. The only treatment for celiac disease is strict, lifelong adherence to a gluten-free diet, which is effective but costly and difficult to maintain. Therefore, novel non-dietary therapies for celiac disease are urgently needed. Gluten-degrading enzymes are promising non-dietary treatments, and some enzymes have been investigated in preclinical or clinical studies. A combination of prolyl endopeptidase from Sphingomonas capsulata (SC PEP) and a glutamine-specific endoprotease (EP-B2 from barley) known as latiglutenase showed insufficient benefits in phase II clinical trials, likely because of its low enzyme activity in the gastric environment. Therefore, improving enzyme activity is essential for the clinical application of SC PEP. Enzyme activity can be enhanced using computer-aided rational protein design tools. In this study, we combined molecular docking and molecular dynamics simulation to rationally design SC PEP mutants and experimentally evaluated their activities. We identified mutants with up to 90–103% increases in specific activity and up to 80–202% increases in the catalytic rate. We have investigated the mechanism underlying the enhanced activity of these mutants, and found that a conformational transition of the β-propeller domain and catalytic domain of SC PEP was important for enzyme activity, and this transition was affected by residues in the catalytic domain and at the domain interface; a shorter distance between the substrate Pro and the oxyanion holes was also crucial for improving SC PEP catalytic activity. Our results provide useful information for the rational design of highly active SC PEPs to accelerate the development of enzyme therapeutics candidates for Celiac disease.