Hydrophilic Residues Research Articles

The Iodide Transport Defect-Causing Y348D Mutation in the Na+/I- Symporter Renders the Protein Intrinsically Inactive and Impairs Its Targeting to the Plasma Membrane.

Background: The sodium/iodide (Na+/I-) symporter (NIS) mediates active transport of I- into the thyroid gland. Mutations in the SLC5A5 gene, which encodes NIS, cause I- transport defects (ITDs)-which, if left untreated, lead to congenital hypothyroidism and consequent cognitive and developmental deficiencies. The ITD-causing NIS mutation Y348D, located in transmembrane segment (TMS) 9, was reported in three Sudanese patients. Methods: We generated cDNAs coding for Y348D NIS and mutants with other hydrophilic and hydrophobic amino acid substitutions at position 348 and transfected them into cells. The activity of the resulting mutants was quantitated by radioiodide transport assays. NIS glycosylation was investigated by Western blotting after endoglycosidase H (Endo H) and PNGase-F glycosidase treatment. Subcellular localization of the mutant proteins was ascertained by flow cytometry analysis, cell surface biotinylation, and immunofluorescence. The intrinsic activity of Y348D was studied by measuring radioiodide transport in membrane vesicles prepared from Y348D-NIS-expressing cells. Our NIS homology models and molecular dynamics simulations were used to identify residues that interact with Y348 and investigate possible interactions between Y348 and the membrane. The sequences of several Slc5 family transporters were aligned, and a phylogenetic tree was generated in ClustalX. Results: Cells expressing Y348D NIS transport no I-. Furthermore, Y348D NIS is only partially glycosylated, is retained intracellularly, and is intrinsically inactive. Hydrophilic residues other than Asp at position 348 also yield NIS proteins that fail to be targeted to the plasma membrane (PM), whereas hydrophobic residues at this position, which we show do not interact with the membrane, rescue PM targeting and function. Conclusions: Y348D NIS does not reach the PM and is intrinsically inactive. Hydrophobic amino acid substitutions at position 348, however, preserve NIS activity. Our findings are consistent with our homology model's prediction that Y348 should face the side opposite the TMS9 residues that coordinate Na+ and participate in Na+ transport, and with the notion that Y348 interacts only with hydrophobic residues. Hydrophilic or charged residues at position 348 have deleterious effects on NIS PM targeting and activity, whereas a hydrophobic residue at this position rescues NIS activity.

Unexpected Role of Achiral Glycine in Determining the Suprastructural Handedness of Peptide Nanofibrils.

Helical supramolecular architectures play important structural and functional roles in biological systems. Although their occurrence is widely perceived to correlate to fundamental chiral units including l-amino acids and d-sugars, the detailed relationship between molecular and supramolecular handedness is still unclear. At the same time, although achiral units are practically always in close proximity to chiral ones by covalent linkage along a polymeric chain, their effect on supramolecular handedness has received relatively less attention. Here, we designed a set of short amphiphilic peptides, in which an achiral glycine residue was incorporated at the interface between the hydrophobic and hydrophilic segments. We observed that glycine incorporation caused dramatic variations in suprastructural handedness in self-assembled peptide nanofibrils, and the effect of the hydrophilic charged residue at the C-terminus on supramolecular handedness was demolished, leading to chiral truncation. Furthermore, molecular dynamics simulations and quantum chemistry calculations revealed that the unanticipated role of the glycine residue in regulating supramolecular handedness originated from its effect on the conformational preference of single β-strands. Importantly, reduced density gradient analyses on single β-strands indicated that, due to the lack of a side chain in glycine, intricate noncovalent interactions were produced among the neighboring amino acid side chains of the incorporated glycine and its local backbone, resulting in diverse β-strand conformations.

Surface Modification of Poly(l-lactic acid) through Stereocomplexation with Enantiomeric Poly(d-lactic acid) and Its Copolymer.

Poly(l-lactic acid) with high molecular weight was used to prepare PLLA films by means of the solvent casting technique. Poly(d-lactic acid) (PDLA) and poly(d-lactic acid-co-glucose) copolymer (PDLAG) with a low molecular weight were synthesized from d-lactic acid and glucose through melt polycondensation. PLLA films were immersed in PDLA or PDLAG solution to prepare surface-modified PLLA films. The modified PLLA film presented stereocomplex crystal (SC) on its surface and homogeneous crystals (HC) in its bulk. The HC structure and surface morphology of modified PLLA films were obviously damaged by PDLA or PDLAG solution. With increasing immersion time, the PLLA films modified by PDLA decreased both the HC and SC structure, while the PLLA films modified by PDLAG increased the SC structure and decreased the HC structure. Hydrophilic glucose residues of PDLAG on the surface would improve the hydrophilicity of surface-modified PLLA films. Moreover, the hydrophilicity of glucose residues and the interaction of glucose residues with lactic acid units could retard HC destruction and SC crystallization, so that PLLA films modified by PDLAG possessed lower melting temperatures of HC and SC, the crystallinity of SC and the water contact angle, compared with PDLAG-modified PLLA films. The SC structure could improve the heat resistance of modified PLLA film, but glucose residues could block crystallization to promote the thermal degradation of PLA materials. The surface modification of PLLA films will improve the thermal stability, hydrophilicity and crystallization properties of PLA materials, which is essential in order to obtain PLA-based biomaterials.

Uniting features from strongly activating small molecules to develop chimeric LRH‐1 agonist

Nuclear receptors are transcription factors that respond to external lipophilic signaling molecules to alter target gene expression. Liver receptor homolog-1 (LRH-1) is a nuclear receptor that operates as a master metabolic regulator in the liver, making it an attractive target for treating a variety of diseases characterized by dysregulated glucose and lipid homeostasis. Phospholipids (PLs) represent the putative endogenous ligands for LRH-1, but their insolubility, rapid turnover, and poor potency have made synthetic agonists more attractive tools for targeting this receptor. With structural data that defined the binding pose of the first class of synthetic LRH-1 agonists, we designed and characterized small molecules with enhanced solubility, potency, and efficacy. This work led to a compound (“6N”) that binds hydrophilic residues deep within the binding pocket, exhibiting nanomolar potency. We have also targeted residues contacted by the headgroups of activating PLs as a complementary strategy for activation. These modifications resulted in our most efficacious small molecule (“10CA”). To capture the characteristics of both compounds, we have synthesized a chimeric small molecule that includes the key moieties of 6N and 10CA responsible for high affinity and efficacy, respectively. Excitingly, this new hybrid compound exhibits the highest affinity, potency, and effect on receptor thermal stability. While the hybrid activates LRH-1 to an equivalent level as 10CA in reporter assays, it elicits LRH-1 target gene expression to a greater degree than either 6N or 10CA alone. Interestingly, RNA-seq studies have revealed that the hybrid compound captures a PL-mediated gene expression response more effectively that either 6N or 10CA alone. Further crystallography and molecular dynamics studies reveal that these characteristics are the result of the hybrid compound engaging with, and communicating from, both sites of agonist binding. Altogether, our studies demonstrate the differences between our leading agonists and suggest that the hybrid molecule is our most effective agonist to date, providing a critical tool for both studying LRH-1 biology and treating a range of metabolic disorders.

Current Understanding of the Structure, Stability and Dynamic Properties of Amyloid Fibrils.

Amyloid fibrils are supramolecular protein assemblies represented by a cross-β structure and fibrous morphology, whose structural architecture has been previously investigated. While amyloid fibrils are basically a main-chain-dominated structure consisting of a backbone of hydrogen bonds, side-chain interactions also play an important role in determining their detailed structures and physicochemical properties. In amyloid fibrils comprising short peptide segments, a steric zipper where a pair of β-sheets with side chains interdigitate tightly is found as a fundamental motif. In amyloid fibrils comprising longer polypeptides, each polypeptide chain folds into a planar structure composed of several β-strands linked by turns or loops, and the steric zippers are formed locally to stabilize the structure. Multiple segments capable of forming steric zippers are contained within a single protein molecule in many cases, and polymorphism appears as a result of the diverse regions and counterparts of the steric zippers. Furthermore, the β-solenoid structure, where the polypeptide chain folds in a solenoid shape with side chains packed inside, is recognized as another important amyloid motif. While side-chain interactions are primarily achieved by non-polar residues in disease-related amyloid fibrils, the participation of hydrophilic and charged residues is prominent in functional amyloids, which often leads to spatiotemporally controlled fibrillation, high reversibility, and the formation of labile amyloids with kinked backbone topology. Achieving precise control of the side-chain interactions within amyloid structures will open up a new horizon for designing useful amyloid-based nanomaterials.

An odorant-binding protein of Asian citrus psyllid, Diaphorina citri, participates in the response of host plant volatiles.

Odorant-binding proteins (OBPs) in insects contribute to the sensitivity of the olfactory system and connect external odorants to olfactory receptor neurons. Determination of the chemosensory functions in Diaphorina citri, a vector of the citrus Huanglongbing pathogen, may help in developing a potential target for pest management. Diaphorina citri showed dose-dependent electroantennogram recording (EAG) responses to 12 host plant volatiles. A two-choice behavioral trap experiment showed that four compounds (methyl salicylate, linalool, citral and R-(+)-limonene) that elicited high EAG responses also had significant attraction to adults. The expression profiles induced by these compounds were detected in nine OBP genes, DcitOBP1-9. DcitOBP3, DcitOBP6 and DcitOBP7 commonly showed significant upregulation or downregulation compared with the control. Microscale thermophoresis (MST) showed that the recombinant protein DcitOBP7 had high in vitro binding affinities (Kd < 10μm) to methyl salicylate, linalool and R-(+)-limonene, and moderate binding affinity to citral with a Kd value of 15.95 μm. Furthermore, RNA interference (RNAi)-suppressed messenger RNA (mRNA) expression of DcitOBP7 resulted in a significant reduction in EAG activity and in adult D. citri behavioral responses to tested volatiles and the preferred host, Murraya paniculata. The hydrophilic residue Arg107 of DcitOBP7 may have a key role in binding odorants via formation of hydrogen bonds. These results show that DcitOBP7 plays an important role in the olfactory response. This finding may provide new insight into the functions of OBP families in D. citri and aid in the development of safe strategies for managing D. citri populations. © 2021 Society of Chemical Industry.

A Reconstructed Common Ancestor of the Fatty Acid Photo-decarboxylase Clade Shows Photo-decarboxylation Activity and Increased Thermostability.

Light‐dependent enzymes are a rare type of biocatalyst with high potential for research and biotechnology. A recently discovered fatty acid photo‐decarboxylase from Chlorella variabilis NC64A (CvFAP) converts fatty acids to the corresponding hydrocarbons only when irradiated with blue light (400 to 520 nm). To expand the available catalytic diversity for fatty acid decarboxylation, we reconstructed possible ancestral decarboxylases from a set of 12 extant sequences that were classified under the fatty acid decarboxylases clade within the glucose‐methanol choline (GMC) oxidoreductase family. One of the resurrected enzymes (ANC1) showed activity in the decarboxylation of fatty acids, showing that the clade indeed contains several photo‐decarboxylases. ANC1 has a 15 °C higher melting temperature (T m) than the extant CvFAP. Its production yielded 12‐fold more protein than this wild type decarboxylase, which offers practical advantages for the biochemical investigation of this photoenzyme. Homology modelling revealed amino acid substitutions to more hydrophilic residues at the surface and shorter flexible loops compared to the wild type. Using ancestral sequence reconstruction, we have expanded the existing pool of confirmed fatty acid photo‐decarboxylases, providing access to a more robust catalyst for further development via directed evolution.

Rational design of Shewanella sp. l-arabinose isomerase for d-galactose isomerase activity under mesophilic conditions

d-Tagatose, a potential low calorific substitute for sucrose, can be produced by bioconversion of d-galactose catalysed by l-arabinose isomerase. l-Arabinose isomerase from Shewanella sp. ANA-3 is unique for its ability to catalyse bioconversion reactions under mesophilic conditions. However, d-galactose not being a natural substrate for l-arabinose isomerase is catalysed at a slower rate. We attempted to increase the biocatalytic efficiency of Shewanella sp. l-arabinose isomerase by rational design to enhance galactose isomerisation activity. In silico molecular docking, analysis has revealed that F279 is sterically hindering the binding of d-galactose at the C6 position. Substitution of bulky Phe residue with smaller hydrophilic residues such as Asn and Thr increased the galactose isomerase activity by 86 % and 12 % respectively. At mesophilic conditions, F279N mutant catalysed the bioconversion of d-galactose more efficiently than l-arabinose, indicating a shift in substrate preference.

Aromatic Rings as Molecular Determinants for the Molecular Recognition of Protein Kinase Inhibitors.

Protein kinases are key enzymes in many signal transduction pathways, and play a crucial role in cellular proliferation, differentiation, and various cell regulatory processes. However, aberrant function of kinases has been associated with cancers and many other diseases. Consequently, competitive inhibition of the ATP binding site of protein kinases has emerged as an effective means of curing these diseases. Over the past three decades, thousands of protein kinase inhibitors (PKIs) with varying molecular frames have been developed. Large-scale data mining of the Protein Data Bank resulted in a database of 2139 non-redundant high-resolution X-ray crystal structures of PKIs bound to protein kinases. This provided us with a unique opportunity to study molecular determinants for the molecular recognition of PKIs. A chemoinformatic analysis of 2139 PKIs resulted in findings that PKIs are “flat” molecules with high aromatic ring counts and low fractions of sp3 carbon. All but one PKI possessed one or more aromatic rings. More importantly, it was found that the average weighted hydrogen bond count is inversely proportional to the number of aromatic rings. Based on this linear relationship, we put forward the exchange rule of hydrogen bonding interactions and non-bonded π-interactions. Specifically, a loss of binding affinity caused by a decrease in hydrogen bonding interactions is compensated by a gain in binding affinity acquired by an increase in aromatic ring-originated non-bonded interactions (i.e., π–π stacking interactions, CH–π interactions, cation–π interactions, etc.), and vice versa. The very existence of this inverse relationship strongly suggests that both hydrogen bonding and aromatic ring-originated non-bonded interactions are responsible for the molecular recognition of PKIs. As an illustration, two representative PKI–kinase complexes were employed to examine the relative importance of different modes of non-bonded interactions for the molecular recognition of PKIs. For this purpose, two FDA-approved PKI drugs, ibrutinib and lenvatinib, were chosen. The binding pockets of both PKIs were thoroughly examined to identify all non-bonded intermolecular interactions. Subsequently, the strengths of interaction energies between ibrutinib and its interacting residues in tyrosine kinase BTK were quantified by means of the double hybrid DFT method B2PLYP. The resulting energetics for the binding of ibrutinib in tyrosine kinase BTK showed that CH–π interactions and π–π stacking interactions between aromatic rings of the drug and hydrophobic residues in its binding pocket dominate the binding interactions. Thus, this work establishes that, in addition to hydrogen bonding, aromatic rings function as important molecular determinants for the molecular recognition of PKIs. In conclusion, our findings support the following pharmacophore model for ATP-competitive kinase inhibitors: a small molecule features a scaffold of one or more aromatic rings which is linked with one or more hydrophilic functional groups. The former has the structural role of acting as a scaffold and the functional role of participating in aromatic ring-originated non-bonded interactions with multiple hydrophobic regions in the ATP binding pocket of kinases. The latter ensure water solubility and form hydrogen bonds with the hinge region and other hydrophilic residues of the ATP binding pocket.

Interaction energy between neuronal cell receptors and peptoid ligands

Peptoids as an extracellular matrix (ECM) material is gaining importance in in vitro neuronal cell culture studies due to their biocompatibility, self-assembling structure, and stability. Mechanotransduction between a neuronal cell and an ECM is mediated by neuronal cell receptors such as integrin and neural cellular adhesion molecule. In this study, using molecular dynamics, we investigate the interaction energies between peptoid and neuronal cell receptors, and also study the effect of peptoid bundle size. We investigate the interaction surface between peptoid bundles and neuronal cell receptors, integrin and neural cellular adhesion molecule, using the solvent accessible surface area method to find the influence of hydrophobic and hydrophilic residues of the peptoid chain. We find the free energy landscape using the umbrella sampling method and then evaluate the potential mean force (PMF) and unbinding force during the dissociation between peptoid bundles and neuronal cell receptors. We find that the peptoid bundles have a higher affinity for the neuronal cell receptors, however increasing the size of peptoid bundles increases the affinity for integrin and neural cell adhesion molecule. PMF data for peptoid and neuronal cell receptor dissociation indicates that binding force increases as the size of the peptoid bundle increases. The higher binding strength during peptoid and neuronal cell receptors are due to the hydrophobic residue cluster area in the binding region. These findings will provide a better insight into using peptoid as an ECM.

Orientational Ambiguity in Septin Coiled Coils and its Structural Basis

Septins are an example of subtle molecular recognition whereby different paralogues must correctly assemble into functional filaments important for essential cellular events such as cytokinesis. Most possess C-terminal domains capable of forming coiled coils which are believed to be involved in filament formation and bundling. Here, we report an integrated structural approach which aims to unravel their architectural diversity and in so doing provide direct structural information for the coiled-coil regions of five human septins. Unexpectedly, we encounter dimeric structures presenting both parallel and antiparallel arrangements which are in consonance with molecular modelling suggesting that both are energetically accessible. These sequences therefore code for two metastable states of different orientations which employ different but overlapping interfaces. The antiparallel structures present a mixed coiled-coil interface, one side of which is dominated by a continuous chain of core hydrophilic residues. This unusual type of coiled coil could be used to expand the toolkit currently available to the protein engineer for the design of previously unforeseen coiled-coil based assemblies. Within a physiological context, our data provide the first atomic details related to the assumption that the parallel orientation is likely formed between septin monomers from the same filament whilst antiparallelism may participate in the widely described interfilament cross bridges necessary for higher order structures and thereby septin function.

A novel insight into transcriptional and epigenetic regulation underlying sex expression and flower development in melon (Cucumis melo L.).

Melon (Cucumis melo L.) is an important cucurbit and has been considered as a model plant for studying sex determination. The four most common sexual morphotypes in melon are monoecious (A-G-M), gynoecious (--ggM-), andromonoecious (A-G-mm), and hermaphrodite (--ggmm). Sex expression in melons is complex, as the genes and associated networks that govern the sex expression are not fully explored. Recently, RNA-seq transcriptomic profiling, ChIP-qPCR analysis integrated with gene ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathways predicted the differentially expressed genes including sex-specific ACS and ACO genes, in regulating the sex-expression, phytohormonal cross-talk, signal transduction, and secondary metabolism in melons. Integration of transcriptional control through genetic interaction in between the ACS7, ACS11, and WIP1 in epistatic or hypostatic manner, along with the recruitment of H3K9ac and H3K27me3, epigenetically, overall determine sex expression. Alignment of protein sequences for establishing phylogenetic evolution, motif comparison, and protein-protein interaction supported the structural conservation while presence of the conserved hydrophilic and charged residues across the diverged evolutionary group predicted the functional conservation of the ACS protein. Presence of the putative cis-binding elements or DNA motifs, and its further comparison with DAP-seq-based cistrome and epicistrome of Arabidopsis, unraveled strong ancestry of melons with Arabidopsis. Motif comparison analysis also characterized putative genes and transcription factors involved in ethylene biosynthesis, signal transduction, and hormonal cross-talk related to sex expression. Overall, we have comprehensively reviewed research findings for a deeper insight into transcriptional and epigenetic regulation of sex expression and flower development in melons.

Promotion and Inhibition of Amyloid-β Peptide Aggregation: Molecular Dynamics Studies.

Aggregates of amyloid-β (Aβ) peptides are known to be related to Alzheimer’s disease. Their aggregation is enhanced at hydrophilic–hydrophobic interfaces, such as a cell membrane surface and air-water interface, and is inhibited by polyphenols, such as myricetin and rosmarinic acid. We review molecular dynamics (MD) simulation approaches of a full-length Aβ peptide, Aβ40, and Aβ(16–22) fragments in these environments. Since these peptides have both hydrophilic and hydrophobic amino acid residues, they tend to exist at the interfaces. The high concentration of the peptides accelerates the aggregation there. In addition, Aβ40 forms a β-hairpin structure, and this structure accelerates the aggregation. We also describe the inhibition mechanism of the Aβ(16–22) aggregation by polyphenols. The aggregation of Aβ(16–22) fragments is caused mainly by the electrostatic attraction between charged amino acid residues known as Lys16 and Glu22. Since polyphenols form hydrogen bonds between their hydroxy and carboxyl groups and these charged amino acid residues, they inhibit the aggregation.

High-resolution structural profile of hylaseptin-4: Aggregation, membrane topology and pH dependence of overall membrane binding process

Hylaseptin-4 (HSP-4, GIGDILKNLAKAAGKAALHAVGESL-NH2) is an antimicrobial peptide originally isolated from Hypsiboas punctatus tree frog. The peptide has been chemically synthetized for structural investigations by CD and NMR spectroscopies. CD experiments reveal the high helical content of HSP-4 in biomimetic media. Interestingly, the aggregation process seems to occur at high peptide concentrations either in aqueous solution or in presence of biomimetic membranes, indicating an increase in the propensity of the peptide for adopting a helical conformation. High-resolution NMR structures determined in presence of DPC-d38 micelles show a highly ordered α-helix from amino acid residues I2 to S24 and a smooth bend near G14. A large separation between hydrophobic and hydrophilic residues occurs up to the A16 residue, from which a shift in the amphipathicity is noticed. Oriented solid-state NMR spectroscopy show a roughly parallel orientation of the helical structure along the POPC lipid bilayer surface, with an insertion of the hydrophobic N-terminus into the bilayer core. Moreover, a noticeable pH dependence of the aggregation process in both aqueous and in biomimetic membrane environments is attributed to a single histidine residue (H19). The protonation degree of the imidazole side-chain might help in modulating the peptide-peptide or peptide-lipid interactions. Finally, molecular dynamics simulations confirm the orientation and preferential helical conformation and in addition, show that HSP-4 tends to self-aggregate in order to stabilize its active conformation in aqueous or phospholipid bilayer environments.

Machine Learning Applied to Determine the Molecular Descriptors Responsible for the Viscosity Behavior of Concentrated Therapeutic Antibodies.

Predicting the solution viscosity of monoclonal antibody (mAb) drug products remains as one of the main challenges in antibody drug design, manufacturing, and delivery. In this work, the concentration-dependent solution viscosity of 27 FDA-approved mAbs was measured at pH 6.0 in 10 mM histidine-HCl. Six mAbs exhibited high viscosity (>30 cP) in solutions at 150 mg/mL mAb concentration. Combining molecular modeling and machine learning feature selection, we found that the net charge in the mAbs and the amino acid composition in the Fv region are key features which govern the viscosity behavior. For mAbs whose behavior was not dominated by charge effects, we observed that high viscosity is correlated with more hydrophilic and fewer hydrophobic residues in the Fv region. A predictive model based on the net charges of mAbs and a high viscosity index is presented as a fast screening tool for classifying low- and high-viscosity mAbs.

DRACP: a novel method for identification of anticancer peptides

BackgroundMillions of people are suffering from cancers, but accurate early diagnosis and effective treatment are still tough for all doctors. Common ways against cancer include surgical operation, radiotherapy and chemotherapy. However, they are all very harmful for patients. Recently, the anticancer peptides (ACPs) have been discovered to be a potential way to treat cancer. Since ACPs are natural biologics, they are safer than other methods. However, the experimental technology is an expensive way to find ACPs so we purpose a new machine learning method to identify the ACPs.ResultsFirstly, we extracted the feature of ACPs in two aspects: sequence and chemical characteristics of amino acids. For sequence, average 20 amino acids composition was extracted. For chemical characteristics, we classified amino acids into six groups based on the patterns of hydrophobic and hydrophilic residues. Then, deep belief network has been used to encode the features of ACPs. Finally, we purposed Random Relevance Vector Machines to identify the true ACPs. We call this method ‘DRACP’ and tested the performance of it on two independent datasets. Its AUC and AUPR are higher than 0.9 in both datasets.ConclusionWe developed a novel method named ‘DRACP’ and compared it with some traditional methods. The cross-validation results showed its effectiveness in identifying ACPs.

Molecular Dynamics Study of Structure, Folding, and Aggregation of Poly-PR and Poly-GR Proteins

Poly-proline-arginine (poly-PR) and poly-glycine-arginine (poly-GR) proteins are believed to be the most toxic dipeptide repeat (DPR) proteins that are expressed by the hexanucleotide repeat expansion mutation in C9ORF72, which are associated with amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) diseases. Their structural information and mechanisms of toxicity remain incomplete, however. Using molecular dynamics simulation and all-atom model of proteins, we study folding and aggregation of both poly-PR and poly-GR. The results indicate formation of double-helix structure during the aggregation of poly-PR into dimers, whereas no stable aggregate is formed during the aggregation of poly-GR; the latter only folds into α-helix and double-helix structures that are similar to those formed in the folding of poly-glycine-alanine (poly-GA) protein. Our findings are consistent with the experimental data indicating that poly-PR and poly-GR are less likely to aggregate because of the hydrophilic arginine residues within their structures. Such characteristics could, however, in some respect facilitate migration of the DPR proteins between and within cells and, at the same time, give proline residues the benefits of activating the receptors that regulate ionotropic effect in neurons, resulting in death or malfunction of neurons because of the abnormal increase or decrease of the ion transmission. This may explain the neurotoxicities of poly-PR and poly-GR associated with many neurodegenerative diseases. To our knowledge, this is the first molecular dynamics simulation of the phenomena involving poly-PR and poly-GR proteins.

Systematically Studying the Optimal Amino Acid Distribution Patterns of the Amphiphilic Structure by Using the Ultrashort Amphiphiles.

Amphipathicity has traditionally been considered to be essential for the de novo design or systematic optimization of antimicrobial peptides (AMPs). However, the current research methods to study the relationship between amphiphilicity and antimicrobial activity are inappropriate, because the key parameters (hydrophobicity, positive charge, etc.) and secondary structure of AMPs are changed. To systematically and accurately study the effects of amphiphilicity on antimicrobial properties of AMPs, we designed parallel series of AMPs with a different order of amino acids in a sequence composed only of Arg and either Trp (WR series) or Leu (LR series), under conditions in which other vital parameters were fixed. Furthermore, based on the WR and LR peptides that can form stable amphiphilic β-sheet structures in the anionic membrane-mimetic environment, we found that high β-sheet amphipathic was accompanied by strong antimicrobial activity. Of such peptides, W5 ([RW]4W) and L5 ([RL]4L) with a nicely amphipathic β-sheet structure possessed the optimal therapeutic index. W5 and L5 also exhibited high stability in vitro and a potent membrane-disruptive mechanism. These results suggest that the alternate arrangement of hydrophobic and hydrophilic residues to form a stable amphipathic β-sheet structure is an essential factor that significantly affects the antimicrobial properties.

A Novel Variant in the TBC1D24 Lipid-Binding Pocket Causes Autosomal Dominant Hearing Loss: Evidence for a Genotype-Phenotype Correlation.

Background: Hereditary hearing loss is a disorder with high genetic and allelic heterogeneity. Diagnostic screening of candidate genes commonly yields novel variants of unknown clinical significance. TBC1D24 is a pleiotropic gene associated with recessive DOORS syndrome, epileptic encephalopathy, myoclonic epilepsy, and both recessive and dominant hearing impairment. Genotype-phenotype correlations have not been established to date but could facilitate diagnostic variant assessment and elucidation of pathomechanisms.Methods and Results: Whole-exome and gene panel screening identified a novel (c.919A>C; p.Asn307His) causative variant in TBC1D24 in two unrelated Caucasian families with Autosomal dominant (AD) nonsyndromic late-onset hearing loss. Protein modeling on the Drosophila TBC1D24 ortholog Skywalker crystal structure showed close interhelix proximity (6.8Å) between the highly conserved residue p.Asn307 in α18 and the position of the single known pathogenic dominant variation (p.Ser178Leu) in α11 that causes a form of deafness with similar clinical characteristics.Conclusion: Genetic variants affecting two polar hydrophilic residues in neighboring helices of TBC1D24 cause AD nonsyndromic late-onset hearing loss. The spatial proximity of the affected residues suggests the first genotype-phenotype association in TBC1D24-related disorders. Three conserved residues in α18 contribute to the formation of a functionally relevant cationic phosphoinositide binding pocket that regulates synaptic vesicle trafficking which may be involved in the molecular mechanism of disease.

Preparation and characterization of genipin cross-linked and lysozyme incorporated antimicrobial sodium caseinate edible films

Sodium caseinate (NaCas) possesses good film forming capacity, but mechanical strength of NaCas films is declined by hydrophilic residues in NaCas molecules. Genipin (GP), a low-toxicity natural cross-linking agent, was used at the level at 1 mg/g NaCas to improve the mechnical properties and lysozyme (LYS) was used at the levels of 42, 84 and 126 mg/g NaCas to confer antimicrobial activity of NaCas films. While films without GP and LYS were colorless, light blueness was observed for all types of NaCas films containing GP. The whiteness index and transparency of films decreased as LYS amount increased. The moisture content of films was not significanly affected by the additon of GP and LYS, but the water vapor permeability slightly increased as the amount of LYS increased. Compared with films without GP, significanly higher tensile strength and Young’s modulus and significantly lower break elongation were detected in NaCas films containing GP. The cross-linking reaction between NaCas and GP was confirmed by FTIR-ATR analysis. Incorporation of LYS into NaCas films exerted antimicobial activities against Staphylococcus aureus and Eschericha coli. In conclusion, a novel type of active edible food packaging film with antimicrobial activity was developed by cross-linked with GP and incoporated with LYS.