Using a dual immunoinformatics and bioinformatics approach to design a novel and effective multi-epitope vaccine against human torovirus disease.

Human Torovirus (HToV), a member of the Coronaviridae family, causes severe enteric diseases with no specific medication available. To develop novel preventative measures, we employed immunoinformatics techniques to design a multi-epitope-based subunit vaccine (HToV-MEV) triggering diverse immune responses. We selected non-allergenic, non-toxic, and antigenic epitopes from structural polyproteins, joined them with suitable linkers, and added an adjuvant 50S ribosomal L7/L12 peptide. The vaccine's solubility score of 0.903678 and physiochemical properties were found effective. Molecular dynamics simulations and free energy calculations revealed strong binding affinity for Toll-like receptor 3 (TLR-3), with a lowest energy score of -303.88, indicating high affinity. In-silico cloning and codon optimization showed significant production potential in E. coli. Immune simulations predicted a human immunological response. Our results are promising, but subsequent in vivo research is recommended. The HToV-MEV vaccine design demonstrates potential for preventing HToV-related diseases, and further investigation is warranted to explore its therapeutic applications.

Promising anti-inflammatory activity of a novel designed anti-microbial peptide for wound healing.

Chronic wounds can develop as a result of prolonged inflammation during the healing process, which can happen due to bacterial infection. Therefore, preventing infection and controlling inflammation can accelerate wound healing. Antimicrobial peptides have different protective properties in addition to antimicrobial activity. Some of these activities include the stimulation of cytokine or chemokine synthesis, the facilitation of chemotaxis and cell proliferation, the acceleration of cell proliferation, the induction of anti-inflammatory responses, and the promotion of wound repair. This study aimed to assess the wound healing potential of a novel in silico-designed antimicrobial peptide. Then, its anti-inflammatory activity was investigated by measuring the level of tumor necrosis factor-α (TNF-α) and transforming growth factor beta (TGF-ß) as indicators of the wound healing process. In addition, the influence of the peptide on cell migration was evaluated by a scratch test on human dermal fibroblasts (HDF) and HaCaT cells as a human epidermal keratinocyte cell line. The results showed that our new peptide could act well in inhibiting TNF-α over-secretion while increasing the expression of TGF-ß as an anti-inflammatory factor. This peptide showed a significant potential to stimulate HDF and HaCaT cell migration and proliferation. Therefore, using this peptide as an anti-inflammatory component of wound dressings may be promising.

Hybrid Caffeic Acid-Based DHFR Inhibitors as Novel Antimicrobial and Anticancer Agents.

A novel series of 1,2,4-triazole analogues of caffeic acid was designed, synthesized, characterized, and assessed for their capacity to inhibit DHFR, as well as their anticancer and antimicrobial properties. A molecular docking analysis was conducted on DHFR, utilizing PDB IDs 1U72 and 2W9S, aiming to design anticancer and antimicrobial drugs, respectively. Among all the synthesized derivatives, compound CTh7 demonstrated the highest potency as a DHFR inhibitor, with an IC50 value of 0.15 µM. Additionally, it exhibited significant cytotoxic properties, with an IC50 value of 8.53 µM. The molecular docking analysis of the CTh7 compound revealed that it forms strong interactions with key residues of homo sapiens DHFR such as Glu30, Phe34, Tyr121, Ile16, Val115, and Phe31 within the target protein binding site and displayed excellent docking scores and binding energy (-9.9; -70.38 kcal/mol). Additionally, synthesized compounds were screened for antimicrobial properties, revealing significant antimicrobial potential against bacterial strains and moderate effects against fungal strains. Specifically, compound CTh3 exhibited notable antibacterial efficacy against Staphylococcus aureus (MIC = 5 µM). Similarly, compound CTh4 demonstrated significant antibacterial activity against both Escherichia coli and Pseudomonas aeruginosa, with MIC values of 5 µM for each. A docking analysis of the most active antimicrobial compound CTh3 revealed that it forms hydrogen bonds with Thr121 and Asn18, a π-cation bond with Phe92, and a salt bridge with the polar residue Asp27.

Designing High Binding Affinity Peptides for MHC Class I Using MAM: An In Silico Approach.

The availability of extensive MHC-peptide binding data has boosted machine learning-based approaches for predicting binding affinity and identifying binding motifs. These computational tools leverage the wealth of binding data to extract essential features and generate a multitude of potential peptides, thereby significantly reducing the cost and time required for experimental procedures. MAM is one such tool for predicting the MHC-I-peptide binding affinity, extracting binding motifs, and generating new peptides with high affinity. This manuscript provides step-by-step guidance on installing, configuring, and executing MAM while also discussing the best practices when using this tool.

Combining Flexible Region Design and Automatic Design to Enhance the Thermal Stability and Catalytic Efficiency of Leucine Dehydrogenase.

Leucine dehydrogenase (LeuDH, EC 1.4.1.9) can reversibly catalyze the oxidative deamination of l-leucine and some other specific α-amino acids to form the corresponding α-ketoacids. This reaction has great significance in the field of food additives and the pharmaceutical industry. The LeuDH from Exiguobacterium sibiricum (EsLeuDH) has high catalytic efficiency but limited thermal stability, hindering its widespread industrial application. In this study, a mutant N5F/I12L/A352Y of EsLeuDH (referred to as M2) was developed with enhanced thermal stability and catalytic activity through rational modification. The M2 mutant exhibits a half-life at 60 °C (t1/2(60 °C)) of 975.7 min and a specific activity of 69.6 U mg-1, which is 5.4 and 2.1 times higher than those of EsLeuDH, respectively. This research may facilitate the utilization of EsLeuDH at elevated temperatures, enhancing its potential for industrial applications. The findings offer a practical and efficient approach for optimizing LeuDH and other industrial enzymes.

Novel pyrrole based CB2 agonists: New insights on CB2 receptor role in regulating neurotransmitters' tone.

The cannabinoid system is one of the most investigated neuromodulatory systems because of its involvement in multiple pathologies such as cancer, inflammation, and psychiatric diseases. Recently, the CB2 receptor has gained increased attention considering its crucial role in modulating neuroinflammation in several pathological conditions like neurodegenerative diseases. Here we describe the rational design of pyrrole-based analogues, which led to a potent and pharmacokinetically suitable CB2 full agonist particularly effective in improving cognitive functions in a scopolamine-induced amnesia murine model. Therefore, we extended our study by investigating the interconnection between CB2 activation and neurotransmission in this experimental paradigm. To this purpose, we performed a MALDI imaging analysis on mice brains, observing that the administration of our lead compound was able to revert the effect of scopolamine on different neurotransmitter tones, such as acetylcholine, serotonin, and GABA, shedding light on important networks not fully explored, so far.

Peptidomimetics as potent dual SARS-CoV-2 cathepsin-L and main protease inhibitors: In silico design, synthesis and pharmacological characterization.

In this paper we present the design, synthesis, and biological evaluation of a new series of peptidomimetics acting as potent anti-SARS-CoV-2 agents. Starting from our previously described Main Protease (MPro) and Papain Like Protease (PLPro) dual inhibitor, CV11, here we disclose its high inhibitory activity against cathepsin L (CTSL) (IC50 = 19.80 ± 4.44 nM), an emerging target in SARS-CoV-2 infection machinery. An in silico design, inspired by the structure of CV11, led to the development of a library of peptidomimetics showing interesting activities against CTSL and Mpro, allowing us to trace the chemical requirements for the binding to both enzymes. The screening in Vero cells infected with 5 different SARS-CoV-2 variants of concerns, highlighted sub-micromolar activities for most of the synthesized compounds (13, 15, 16, 17 and 31) in agreement with the enzymatic inhibition assays results. The compounds showed lack of activity against several different RNA viruses except for the 229E and OC43 human coronavirus strains, also characterized by a cathepsin-L dependent release into the host cells. The most promising derivatives were also evaluated for their chemical and metabolic in-vitro stability, with derivatives 15 and 17 showing a suitable profile for further preclinical characterization.

In silico design of novel bioactive molecules to treat breast cancer with chlorogenic acid derivatives: a computational and SAR approach.

Introduction: Cancer is a vast group of diseases comprising abnormal cells that multiply and grow uncontrollably, and it is one of the top causes of death globally. Several types of cancers are diagnosed, but the incidence of breast cancer, especially in postmenopausal women, is increasing daily. Chemotherapeutic agents used to treat cancer are generally associated with severe side effects on host cells, which has led to a search for safe and potential alternatives. Therefore, the present research has been conducted to find novel bioactive molecules to treat breast cancer with chlorogenic acid and its derivatives. Chlorogenic acid was selected because of its known activity in the field. Methods: Several chlorogenic acid derivatives were subjected to computational studies such as molecular docking, determination of absorption, distribution, metabolism, and excretion (ADME), druglikeness, toxicity, and prediction of activity spectra for substances (PASS) to develop a potential inhibitor of breast cancer. The Protein Data Bank (PDB) IDs used for docking purposes were 7KCD, 3ERT, 6CHZ, 3HB5, and 1U72. Result: Exhaustive analysis of results has been conducted by considering various parameters, like docking score, binding energy, types of interaction with important amino acid residues in the binding pocket, ADME, and toxicity data of compounds. Among all the selected derivatives, CgE18, CgE11, CgAm13, CgE16, and CgE9 have astonishing interactions, excellent binding energy, and better stability in the active site of targeted proteins. The docking scores of compound CgE18 were -11.63 kcal/mol, -14.15 kcal/mol, and -12.90 kcal/mol against breast cancer PDB IDs 7KCD, 3HB5, and 1U72, respectively. The docking scores of compound CgE11 were -10.77 kcal/mol and -9.11 kcal/mol against breast cancer PDB IDs 3ERT and 6CHZ, respectively, whereas the docking scores of epirubicin hydrochloride were -3.85 kcal/mol, -6.4 kcal/mol, -8.76 kcal/mol, and -10.5 kcal/mol against PDB IDs 7KCD, 3ERT, 6CHZ, and 3HB5. The docking scores of 5-fluorouracil were found to be -5.25 kcal/mol, -3.43 kcal/mol, -3.73 kcal/mol, and -5.29 kcal/mol against PDB IDs 7KCD, 3ERT, 6CHZ, and 3HB5, which indicates the designed compounds have a better docking score than some standard drugs. Conclusion: Taking into account the results of molecular docking, drug likeness analysis, absorption, distribution, metabolism, excretion, and toxicity (ADMET) evaluation, and PASS, it can be concluded that chlorogenic acid derivatives hold promise as potent inhibitors for the treatment of breast cancer.

In Silico-Based Design of a Hybrid Peptide with Antimicrobial Activity against Multidrug-Resistant Pseudomonas aeruginosa Using a Spider Toxin Peptide.

The escalating prevalence of antibiotic-resistant bacteria poses an immediate and grave threat to public health. Antimicrobial peptides (AMPs) have gained significant attention as a promising alternative to conventional antibiotics. Animal venom comprises a diverse array of bioactive compounds, which can be a rich source for identifying new functional peptides. In this study, we identified a toxin peptide, Lycotoxin-Pa1a (Lytx-Pa1a), from the transcriptome of the Pardosa astrigera spider venom gland. To enhance its functional properties, we employed an in silico approach to design a novel hybrid peptide, KFH-Pa1a, by predicting antibacterial and cytotoxic functionalities and incorporating the amino-terminal Cu(II)- and Ni(II) (ATCUN)-binding motif. KFH-Pa1a demonstrated markedly superior antimicrobial efficacy against pathogens, including multidrug-resistant (MDR) Pseudomonas aeruginosa, compared to Lytx-Pa1a. Notably, KFH-Pa1a exerted several distinct mechanisms, including the disruption of the bacterial cytoplasmic membrane, the generation of intracellular ROS, and the cleavage and inhibition of bacterial DNA. Additionally, the hybrid peptide showed synergistic activity when combined with conventional antibiotics. Our research not only identified a novel toxin peptide from spider venom but demonstrated in silico-based design of hybrid AMP with strong antimicrobial activity that can contribute to combating MDR pathogens, broadening the utilization of biological resources by incorporating computational approaches.

Improved production of Taxol® precursors in S. cerevisiae using combinatorial in silico design and metabolic engineering.

BACKGROUND: Integrated metabolic engineering approaches that combine system and synthetic biology tools enable the efficient design of microbial cell factories for synthesizing high-value products. In this study, we utilized in silico design algorithms on the yeast genome-scale model to predict genomic modifications that could enhance the production of early-step Taxol® in engineered Saccharomyces cerevisiae cells. RESULTS: Using constraint-based reconstruction and analysis (COBRA) methods, we narrowed down the solution set of genomic modification candidates. We screened 17 genomic modifications, including nine gene deletions and eight gene overexpressions, through wet-lab studies to determine their impact on taxadiene production, the first metabolite in the Taxol® biosynthetic pathway. Under different cultivation conditions, most single genomic modifications resulted in increased taxadiene production. The strain named KM32, which contained four overexpressed genes (ILV2, TRR1, ADE13, and ECM31) involved in branched-chain amino acid biosynthesis, the thioredoxin system, de novo purine synthesis, and the pantothenate pathway, respectively, exhibited the best performance. KM32 achieved a 50% increase in taxadiene production, reaching 215 mg/L. Furthermore, KM32 produced the highest reported yields of taxa-4(20),11-dien-5α-ol (T5α-ol) at 43.65 mg/L and taxa-4(20),11-dien-5-α-yl acetate (T5αAc) at 26.2 mg/L among early-step Taxol® metabolites in S. cerevisiae. CONCLUSIONS: This study highlights the effectiveness of computational and integrated approaches in identifying promising genomic modifications that can enhance the performance of yeast cell factories. By employing in silico design algorithms and wet-lab screening, we successfully improved taxadiene production in engineered S. cerevisiae strains. The best-performing strain, KM32, achieved substantial increases in taxadiene as well as production of T5α-ol and T5αAc. These findings emphasize the importance of using systematic and integrated strategies to develop efficient yeast cell factories, providing potential implications for the industrial production of high-value isoprenoids like Taxol®.

Engineering of formate dehydrogenase for improving conversion potential of carbon dioxide to formate.

Formate dehydrogenase (FDH) is a D-2-hydroxy acid dehydrogenase, which can reversibly reduce CO2 to formate and thus act as non-photosynthetic CO2 reductase. In order to increase catalytic efficiency of formate dehydrogenase for CO2 reduction, two mutants V328I/F285W and V354G/F285W were obtained of which reduction activity was about two times more than the parent CbFDHM2, and the formate production from CO2 catalyzed by mutants were 2.9 and 2.7-fold higher than that of the parent CbFDHM2. The mutants had greater potential in CO2 reduction. The optimal temperature for V328I/F285W and V354G/F285W was 55 °C, and they showed increasement of relative activity under 45 °C to 55 °C compared with parent. The optimal pH for the mutants was 9.0, and they showed excellent stability in pH 4.0-11.5. The kcat/Km values of mutants were 1.75 times higher than that of the parent. Then the molecular basis for its improvement of biochemical characteristics were preliminarily elucidated by computer-aided methods. All of these results further established a solid foundation for molecular modification of formate dehydrogenase and CO2 reduction.

Molecularly imprinted polymer-based sensors for identification volatile compounds in pharmaceutical products: in silico rational design.

The present study aimed to strategically design a Molecularly Imprinted Polymer (MIP) with selective extraction capabilities for volatile compounds found in pork. These specific volatile compounds, such as 3-methyl-1-butanol, 1-nonanal, octanal, hexanal, 2-pentyl-furan, 1-penten-3-one, N-morpholinomethyl-isopropyl-sulfide, methyl butyrate, and (E,E)-2,4-decadienal, are primarily responsible for the distinctive aroma and flavor characteristics associated with pork. Molecular dynamics simulations were employed to investigate the stability of the pre-polymerization system, simulating the interactions between the volatile compounds as templates, 4-hydroxyethyl methacrylate (HEMA) as monomers, and ethylene glycol dimethacrylate (EGDMA) as crosslinkers. Computational simulations revealed that the optimal mole ratio of 1:4:20 for templates, monomers, and crosslinkers resulted in the most favorable functional radial distribution and exhibited the strongest interactions. To validate the computational findings, additional analyses were performed utilizing Molecular Mechanics Poisson-Boltzmann Surface Area (MM-PBSA), radial distribution function (RDF), and hydrogen bond (HBond) occupancy. The calculated binding free energy demonstrated that all template molecules were capable to bind with both the monomers and crosslinkers, including 1-penten-3-one and N-morpholinomethyl-isopropyl-sulfide displaying the strongest interactions, with values of -12,674 kJ/mol and -11,646 kJ/mol, respectively. The congruence between the results obtained from the molecular simulation analyses highlights the crucial role of molecular dynamics simulations in the study and development of MIP for the analysis of marker compounds present in pork.Communicated by Ramaswamy H. Sarma.

Computer-Assisted Design and Characterization of RNA Nanostructures.

Molecular dynamics (MD) simulations can aid in the design and characterization of RNA nanomaterials, providing details about structural and dynamical properties as a function of sequence and environment. Here, we describe how to perform explicit and implicit solvent all-atom MD simulations for RNA nanoring systems.

Design, in silico and pharmacological evaluation of a peptide inhibitor of BACE-1.

Introduction: Alzheimer's disease (AD) is the main type of dementia, caused by the accumulation of amyloid plaques, formed by amyloid peptides after being processed from amyloid precursor protein (APP) by γ- and ß-secretases (BACE-1). Although amyloid peptides have been well established for AD, they have been found in other neurodegenerative diseases, such as Parkinson's disease, Lewy body dementia, and amyotrophic lateral sclerosis. Inhibitors of BACE-1 have been searched and developed, but clinical trials failed due to lack of efficacy or toxicity. Nevertheless, it is still considered a good therapeutic target, as it was proven to remove amyloid peptides and improve memory. Methods: In this work, we designed a peptide based on a sequence obtained from the marine fish Merluccius productus and evaluated it by molecular docking to verify its binding to BACE-1, which was tested experimentally by enzymatic kinetics and cell culture assays. The peptide was injected in healthy mice to study its pharmacokinetics and toxicity. Results: We could obtain a new sequence in which the first N-terminal amino acids and the last one bound to the catalytic site of BACE-1 and showed high stability and hydrophobicity. The synthetic peptide showed a competitive inhibition of BACE-1 and Ki = 94 nM, and when injected in differentiated neurons, it could reduce Aß42o production. In plasma, its half-life is ∼1 h, clearance is 0.0015 µg/L/h, and Vss is 0.0015 µg/L/h. The peptide was found in the spleen and liver 30 min after injection and reduced its level after that, when it was quantified in the kidneys, indicating its fast distribution and urinary excretion. Interestingly, the peptide was found in the brain 2 h after its administration. Histological analysis showed no morphological alteration in any organ, as well as the absence of inflammatory cells, indicating a lack of toxicity. Discussion: We obtained a new BACE-1 inhibitor peptide with fast distribution to the tissues, without accumulation in any organ, but found in the brain, with the possibility to reach its molecular target, BACE-1, contributing to the reduction in the amyloid peptide, which causes amyloid-linked neurodegenerative diseases.

Improved HIV-1 neutralization breadth and potency of V2-apex antibodies by in silico design.

Broadly neutralizing antibodies (bNAbs) against HIV can reduce viral transmission in humans, but an effective therapeutic will require unusually high breadth and potency of neutralization. We employ the OSPREY computational protein design software to engineer variants of two apex-directed bNAbs, PGT145 and PG9RSH, resulting in increases in potency of over 100-fold against some viruses. The top designed variants improve neutralization breadth from 39% to 54% at clinically relevant concentrations (IC80 < 1 µg/mL) and improve median potency (IC80) by up to 4-fold over a cross-clade panel of 208 strains. To investigate the mechanisms of improvement, we determine cryoelectron microscopy structures of each variant in complex with the HIV envelope trimer. Surprisingly, we find the largest increases in breadth to be a result of optimizing side-chain interactions with highly variable epitope residues. These results provide insight into mechanisms of neutralization breadth and inform strategies for antibody design and improvement.

In silico optimization of peptides that inhibit Wnt/ß-catenin signaling.

The Wnt/ß-catenin signaling pathway causes transcriptional activation through the interaction between ß-catenin and T cell-specific transcription factor (TCF) and regulates a wide variety of cellular responses, including proliferation, differentiation and cell motility. Excessive transcriptional activation of the Wnt/ß-catenin pathway is implicated in developing or exacerbating various cancers. We have recently reported that liver receptor homolog-1 (LRH-1)-derived peptides inhibit the ß-catenin/TCF interaction. In addition, we developed a cell-penetrating peptide (CPP)-conjugated LRH-1-derived peptide that inhibits the growth of colon cancer cells and specifically inhibits the Wnt/ß-catenin pathway. Nonetheless, the inhibitory activity of the CPP-conjugated LRH-1-derived peptide was unsatisfactory (ca. 20 µM), and improving the bioactivity of peptide inhibitors is required for their in vivo applications. In this study, we optimized the LRH-1-derived peptide using in silico design to enhance its activity further. The newly designed peptides showed binding affinity toward ß-catenin comparable to the parent peptide. In addition, the CPP-conjugated stapled peptide, Penetratin-st6, showed excellent inhibition (ca. 5 µM). Thus, the combination of in silico design by MOE and MD calculations has revealed that logical molecular design of PPI inhibitory peptides targeting ß-catenin is possible. This method can be also applied to the rational design of peptide-based inhibitors targeting other proteins.

In Silico Design and Experimental Validation of Novel Oxazole Derivatives Against Varicella zoster virus.

Varicella zoster virus (VZV) infection causes severe disease such as chickenpox, shingles, and postherpetic neuralgia, often leading to disability. Reactivation of latent VZV is associated with a decrease in specific cellular immunity in the elderly and in patients with immunodeficiency. However, due to the limited efficacy of existing therapy and the emergence of antiviral resistance, it has become necessary to develop new and effective antiviral drugs for the treatment of diseases caused by VZV, particularly in the setting of opportunistic infections. The goal of this work is to identify potent oxazole derivatives as anti-VZV agents by machine learning, followed by their synthesis and experimental validation. Predictive QSAR models were developed using the Online Chemical Modeling Environment (OCHEM). Data on compounds exhibiting antiviral activity were collected from the ChEMBL and uploaded in the OCHEM database. The predictive ability of the models was tested by cross-validation, giving coefficient of determination q2 = 0.87-0.9. The validation of the models using an external test set proves that the models can be used to predict the antiviral activity of newly designed and known compounds with reasonable accuracy within the applicability domain (q2 = 0.83-0.84). The models were applied to screen a virtual chemical library with expected activity of compounds against VZV. The 7 most promising oxazole derivatives were identified, synthesized, and tested. Two of them showed activity against the VZV Ellen strain upon primary in vitro antiviral screening. The synthesized compounds may represent an interesting starting point for further development of the oxazole derivatives against VZV. The developed models are available online at OCHEM http://ochem.eu/article/145978 and can be used to virtually screen for potential compounds with anti-VZV activity.

Bioinformatics design of recombinant chimeric protein containing SipD and LptD immunogens and evaluation of its immunogenicity against Salmonella Typhimurium.

The growing emergence of resistant bacteria is the current global concern for the humans and animals. Vaccination could be the desirable method to preventing such infectious diseases. Safe and effective vaccines are urgently needed to manage and prevent Salmonella contamination. Subunit vaccines are safe approaches for the protection against Salmonella spp. The bioinformatics methods were performed to determine the gene structure. Gene cassette (rLPSI) was ordered in pET28a (+), and cloned into E.coli BL21 (DE3), and the recombinant protein was expressed using IPTG (1 mM). Mice were immunized by subcutaneous administration of recombinant protein. Then, the mice were challenged by oral administration of 100LD50 of live S. Typhimurium. The Codon adaptation index of the chimeric gene was multiplied by 0.92. Validation results showed that >90% of residues lie in the desired or extra allowed area of the Ramachandran plot. The recombinant protein (65.9 kDa) was expressed in E.coli. Antibody titers in vaccinated mice were significantly different from those in the control groups. Recombinant protein immunization of the mice provided 90% and 70% protection against 10LD50 and 100LD50 of S. Typhimurium, respectively. In general, the results showed the high efficiency of rLPSI chimeric protein as a protective antigen against S. Typhimurium infection. The rLPSI chimeric protein could be an effective recombinant vaccine candidate against S. Typhimurium infection, but more research is needed.

Design, in silico and pharmacological evaluation of a peptide inhibitor of BACE-1

Introduction: Alzheimer’s disease (AD) is the main type of dementia, caused by the accumulation of amyloid plaques, formed by amyloid peptides after being processed from amyloid precursor protein (APP) by γ- and ß-secretases (BACE-1). Although amyloid peptides have been well established for AD, they have been found in other neurodegenerative diseases, such as Parkinson’s disease, Lewy body dementia, and amyotrophic lateral sclerosis. Inhibitors of BACE-1 have been searched and developed, but clinical trials failed due to lack of efficacy or toxicity. Nevertheless, it is still considered a good therapeutic target, as it was proven to remove amyloid peptides and improve memory. Methods: In this work, we designed a peptide based on a sequence obtained from the marine fish Merluccius productus and evaluated it by molecular docking to verify its binding to BACE-1, which was tested experimentally by enzymatic kinetics and cell culture assays. The peptide was injected in healthy mice to study its pharmacokinetics and toxicity. Results: We could obtain a new sequence in which the first N-terminal amino acids and the last one bound to the catalytic site of BACE-1 and showed high stability and hydrophobicity. The synthetic peptide showed a competitive inhibition of BACE-1 and Ki = 94 nM, and when injected in differentiated neurons, it could reduce Aβ42o production. In plasma, its half-life is ∼1 h, clearance is 0.0015 μg/L/h, and Vss is 0.0015 μg/L/h. The peptide was found in the spleen and liver 30 min after injection and reduced its level after that, when it was quantified in the kidneys, indicating its fast distribution and urinary excretion. Interestingly, the peptide was found in the brain 2 h after its administration. Histological analysis showed no morphological alteration in any organ, as well as the absence of inflammatory cells, indicating a lack of toxicity. Discussion: We obtained a new BACE-1 inhibitor peptide with fast distribution to the tissues, without accumulation in any organ, but found in the brain, with the possibility to reach its molecular target, BACE-1, contributing to the reduction in the amyloid peptide, which causes amyloid-linked neurodegenerative diseases.

Exploring the active core of a novel antimicrobial peptide, palustrin-2LTb, from the Kuatun frog, Hylarana latouchii, using a bioinformatics-directed approach.

Antimicrobial peptides (AMPs), one of the most promising next-generation antibiotics to address the problem of antibiotic-resistance, have gained increasing attention in recent decades. However, some bottlenecks, such as high manufacturing costs and high toxicity, have greatly hindered their development. To overcome these problems, we developed an efficient modification approach to find the valid active-core fragments of AMPs by mimicking the cleavage process of trypsin-like specificity proteases in silico, and truncating the peptide. Herein, we used the structure of a novel AMP, palustrin-2LTb, as the template and synthesised a set of interceptive peptides using computer-aided design and prediction. Functional screening data indicated that truncated fragment 3 not only maintained and optimised antimicrobial efficacy of the parent peptide but also showed great in vivo therapeutic potential in an MRSA-infected insect larvae model. Overall, the demonstration of the therapeutic efficacy of fragment 3 showcases the efficiency of our approach for future modification of AMPs.