Cryptides are a subfamily of bioactive peptides embedded latently in their parent proteins and have multiple biological functions. Cationic cryptides could be used as modern drugs in both infectious diseases and cancers because their mechanism of action is less likely to be affected by genetic mutations in the treated cells, therefore addressing a current unmet need in these two areas of medicine. In this review, we present the current understanding of cryptides, methods to mine them sustainably using available online databases and prediction tools, with a particular focus on their antimicrobial and anticancer potential, and their potential applicability in a clinical setting.
A previous study has shown that synthetic antimicrobial peptides (AMPs) derived from Anabas testudineus (ATMP1) could in-vitro inhibit the progression of breast cancer cell lines. In this study, we are interested in studying altered versions of previous synthetic AMPs to gain some insight into the peptides functions. The AMPs were altered and subjected to bioinformatics prediction using four databases (ADP3, CAMP-R3, AMPfun, and ANTICP) to select the highest anticancer activity. The bioinformatics in silico analysis led to the selection of two AMPs, which are ATMP5 (THPPTTTTTTTTTTTYTAAPATTT) and ATMP6 (THPPTTTTTTTTTTTTTAAPARTT). The in silico analysis predicted that ATMP5 and ATMP6 have anticancer activity and lead to cell death. The ATMP5 and ATMP6 were submitted to deep learning databases (ToxIBTL and ToxinPred2) to predict the toxicity of the peptides and to (AllerTOP & AllergenFP) check the allergenicity. The results of databases indicated that AMPs are non-toxic to normal human cells and allergic to human immunoglobulin. The bioinformatics findings led to select the highest active peptide ATMP5, which was synthesised and applied for in-vitro experiments using cytotoxicity assay MTT Assay, apoptosis detection using the Annexin V FTIC-A assay, and gene expression using Apoptosis PCR Array to evaluate the AMP's anticancer activity. The antimicrobial activity is approved by the disc diffusion method. The in-vitro experiments analysis showed that ATMP5 had the activity to inhibit the growth of the breast cancer cell line (MDA-MB-231) after 48 h and managed to arrest the cell cycle of the MDA-MB-231, apoptosis induction, and overexpression of the p53 by interaction with the related apoptotic genes. This research opened up new opportunities for developing potential and selective anticancer agents relying on antimicrobial peptide properties.
The global pandemic of COVID-19 is a serious public health concern. Over 625 million confirmed cases and more than 6 million deaths have been recorded worldwide. Although several vaccines and antiviral medications have been developed, their efficacy is limited by the emerging new SARS-CoV-2 strains. Peptide-based therapeutics is a fast-growing class of new drugs and have unique advantages over large proteins and small molecules. Antiviral peptides (AVPs) are short polycationic antivirals with broad-spectrum effects, which have been shown to exert both prophylactic and therapeutic actions against reported coronaviruses. The potential therapeutic targets of AVPs are located either on the virus (e.g., E-protein and S-protein) to prohibit viral binding or host cells, particularly, those present on the cell surface (e.g., ACE2 and TMPRSS2). Despite AVPs having promising antiviral effects, their efficacy is limited by low bioavailability. Thus, nanoformulation is a prerequisite for prolonged bioavailability and efficient delivery. This review aimed to present an insight into the therapeutic AVP targets on both virus and host cells by discussing their antiviral activities and associated molecular mechanisms. Besides, it described the technique for discovering and developing possible AVPs based on their targets, as well as the significance of using nanotechnology for their efficient delivery against SARS-CoV-2.
Nowadays dengue virus infection (DENV) is one of the major health complications in the world. Although DENV is an old and common disease, unfortunately, until now, there are no specific relevant treatments available for it. This study, therefore, aimed to design, as well as synthesize selective peptide inhibitors, and investigate their activity by in-vitro NS2B/NS3 protease inhibition assay. The design of the peptide ligands was based on studying the interactions with the dengue NS2B/NS3 protease using the computational docking technique in the MOE and AutoDock (version 4.2) software. To this end, the researchers designed 26 linear pentapeptides based on previous studies. It was revealed that two linear pentapeptides (i.e., GKRRK and KRRRK) are the best potential inhibitors. Furthermore, based on the findings of the two independent docking programs, the peptide GKRRK was synthesized by solid-phase peptide synthesis and its structure was confirmed. The in-vitro protease inhibitor study was conducted for these two peptides to examine their activity against the dengue virus using a protin in as a control. It was found that the designed potential peptides possess interesting inhibition against the NS2B/NS3 protease. Additionally, the findings showed that the peptide GKRRK had the highest percentage of inhibition (71.11%) at 100 µM with the IC50 of 48.87 µM; therefore, this linear peptide could serve as a good inhibitor for the DENV.
Antibiotic resistance is a growing concern that is affecting public health globally. The search for alternative antimicrobial agents has become increasingly important. Antimicrobial peptides (AMPs) produced by Bacillus spp. have emerged as a promising alternative to antibiotics, due to their broad-spectrum antimicrobial activity against resistant pathogens. In this review, we provide an overview of Bacillus-derived AMPs, including their classification into ribosomal (bacteriocins) and non-ribosomal peptides (lipopeptides and polyketides). Additionally, we delve into the molecular mechanisms of AMP production and describe the key biosynthetic gene clusters involved. Despite their potential, the low yield of AMPs produced under normal laboratory conditions remains a challenge to large-scale production. This review thus concludes with a comprehensive summary of recent studies aimed at enhancing the productivity of Bacillus-derived AMPs. In addition to medium optimization and genetic manipulation, various molecular strategies have been explored to increase the production of recombinant antimicrobial peptides (AMPs). These include the selection of appropriate expression systems, the engineering of expression promoters, and metabolic engineering. Bacillus-derived AMPs offer great potential as alternative antimicrobial agents, and this review provides valuable insights on the strategies to enhance their production yield, which may have significant implications for combating antibiotic resistance. KEY POINTS: • Bacillus-derived AMP is a potential alternative therapy for resistant pathogens • Bacillus produces two main classes of AMPs: ribosomal and non-ribosomal peptides • AMP yield can be enhanced using culture optimization and molecular approaches.
The application of proteomic and peptidomic technologies for food-derived bioactive peptides is an emerging field in food sciences. These technologies include the use of separation tools coupled to a high-resolution spectrometric and bioinformatic tools for prediction, identification, sequencing, and characterization of peptides. To a large extent, one-dimensional separation technologies have been extensively used as a continuous tool under different optimized conditions for the identification and analysis of food peptides. However, most one-dimensional separation technologies are fraught with significant bottlenecks such as insufficient sensitivity and specificity limits for complex samples. To address this limitation, separation systems based on orthogonal, multidimensional principles, which allow for the coupling of more than one analytical separation tool with different operational principles, provide a higher separation power than one-dimensional separation tools. This review describes the structure-informed separation and purification of protein hydrolyzates to obtain peptides with desirable bioactivities. PRACTICAL APPLICATIONS: Application of bioactive peptides in the formulation of functional foods, nutraceuticals, and therapeutic agents have increasingly gained scholarly and industrial attention. The bioactive peptides exist originally in protein sources and are only active after hydrolysis of the parent protein. Currently, several tools can be configured in one-dimensional or multidimensional systems for the separation and purification of protein hydrolyzates. The separations are informed by the structural properties such as the molecular weight, charge, hydrophobicity or hydrophilicity, and the solubility of peptides. This review provides a concise discussion on the commonly used analytical tools, their configurations, advantages and challenges in peptide separation. Emphasis is placed on how the structural properties of peptides assist in the separation and purification processes and the concomitant effect of the separation on peptide bioactivity.
Macrobrachium rosenbergii nodavirus (MrNv) causes white tail disease (WTD) in giant freshwater prawns, which leads to devastating economic losses in the aquaculture industry. Despite extensive research on MrNv, there is still no antiviral agent to treat WTD. Thus, the main aim of this study was to identify potential anti-MrNv molecules. A 12-mer phage-displayed peptide library was biopanned against the MrNv virus-like particle (VLP). After four rounds of biopanning, two dominant phages harbouring the amino acid sequences HTKQIPRHIYSA and VSRHQSWHPHDL were selected. An equilibrium binding assay in solution was performed to determine the relative dissociation constant (KDrel) of the interaction between the MrNv VLP and the selected fusion phages. Phage-HTKQIPRHIYSA has a KDrel value of 92.4±22.8 nM, and phage-VSRHQSWHPHDL has a KDrel value of 12.7±3.8 nM. An in-cell elisa was used to determine the inhibitory effect of the synthetic peptides towards the entry of MrNv VLP into Spodoptera frugiperda (Sf9) cells. Peptides HTKQIPRHIYSA and VSRHQSWHPHDL inhibited the entry of the MrNv VLP into Sf9 cells with IC50 values of 30.4±3.6 and 26.5±8.8 µM, respectively. Combination of both peptides showed a significantly higher inhibitory effect with an IC50 of 4.9±0.4 µM. An MTT assay revealed that the viability of MrNv-infected cells increased to about 97 % in the presence of both peptides. A real-time RT-PCR assay showed that simultaneous application of both peptides significantly reduced the number of MrNv per infected cell, from 97±9 to 11±4. These peptides are lead compounds which can be further developed into potent anti-MrNv agents.
This study investigatedthe mechanism of epinecidin-1 against Botrytis cinerea, in vitro, and its effectiveness at inhibiting gray mold on postharvest peach fruit. We found that in vitro, epinecidin-1 had significantly greater antifungal activity against B. cinerea than either clavanin-A or mytimycin, two other marine derived antimicrobial peptides that we tested. Its antifungal activity was heat-resistant (15 min at 40-100 °C) and tolerant to lower concentrations of cations (<100 mM Na+, K+; <10 mM Ca2+). Epinecidin-1 interacted directly with B. cinerea genomic DNA, and that in mycelia, epinecidin-1 exposure induced accumulation of intracellular ROS and increased the permeability of cell membranes resulting in leakage of nucleic acids and aberrant cell morphology. Meanwhile, 200 μM of epinecidin-1 had a significant inhibitory effect on gray mold injected into peach fruit. These results suggested that epinecidin-1 showed promise as a potential method for controlling postharvest gray mold in peaches.
The medicinal Malaysian leeches have been used in traditional medicine to treat many different ailments. In this study, leech saliva extract (LSE) was collected from the medicinal Malaysian leech Hirudinaria manillensis. Gel electrophoresis of LSE was carried out to estimate the peptide and protein molecular weights of its content. Results showed that LSE contains more than 60 peptides and proteins with molecular masses ranging from 1.9-250kDa. Thrombin time assay in vitro was employed to assess the collected LSE antithrombin activity. First, to study its stability, LSE was lyophilized under the following different conditions: pre-freezing temperature, type of container and lyophilization cycle. Pre-freezed LSE sample at -20°C and lyophilized for 24 hours retained about 100-95% of its original biological activities. Second, the LSE antithrombin activity was monitored for a period of six months. Storage temperature, type of the container and photosensitivity effects on antithrombin activity of the lyophilized (solid state) and non-lyophilized (liquid state) were investigated. Results showed that storage temperature drastically affected the biological activity of LSE with -20 °C as the optimum temperature. Samples stored at ambient temperature and +4 °C were light photosensitive and adversely affected when stored in polypropylene tubes. Lyophilized samples were more stable than non-lyophilized ones over the period of study. To sum up, in order to have a biologically active stock of LSE, it has to be lyophilized for no more than 24 hours following freezing at -20°C and has to be stored at -20°C in glass tubes protected from light.
Platelets release more than 30 cytokines to provide primary hemostatic function. In addition, platelets are also known to release antimicrobial peptides upon activation by thrombin.
Fishes are rich sources of structurally diverse bioactive compounds. In recent years, much attention has been paid to the existence of peptides with biological activities and proteins derived from foods that might have beneficial effects for humans. Antioxidant and antimicrobial peptides isolated from fish sources may be used as functional ingredients in food formulations to promote consumer health and improve the shelf life of food products. This paper presents an overview of the antioxidant and antimicrobial peptides derived from various fishes. In addition, we discuss the extraction of fish proteins, enzymatic production, and the techniques used to isolate and characterize these compounds. Furthermore, we review the methods used to assay the bioactivities and their applications in food and nutraceuticals.
Avian influenza viruses (AIV) cause high morbidity and mortality among the poultry worldwide. Their highly mutative nature often results in the emergence of drug resistant strains, which have the potential of causing a pandemic. The virus has two immunologically important glycoproteins, hemagglutinin (HA), neuraminidase (NA), and one ion channel protein M2 which are the most important targets for drug discovery, on its surface. In order to identify a peptide-based virus inhibitor against any of these surface proteins, a disulfide constrained heptapeptide phage display library was biopanned against purified AIV sub-type H9N2 virus particles.
The ability of Leuconostoc mesenteroides DU15 to produce antifungal peptides that inhibit growth of Aspergillus niger was evaluated under optimum growth conditions of 30 °C for 48 h. The cell-free supernatant showed inhibitory activity against A. niger. Five novel peptides were isolated with the sequences GPFPL, YVPLF, LLHGVPLP, GPFPLEMTLGPT, and TVYPFPGPL as identified by de novo sequencing using PEAKS 6 software. Peptide LLHGVPLP was the only positively charged (cationic peptides) and peptide GPFPLEMTLGPT negatively charged (anionic), whereas the rest are neutral. The identified peptides had high hydrophobicity ratio and low molecular weights with amino acids sequences ranging from 5 to 12 residues. The mode of action of these peptides is observed under the scanning electron microscope and is due to cell lysis of fungi. This work reveals the potential of peptides from L. mesenteroides DU15 as natural antifungal preservatives in inhibiting the growth of A. niger that is implicated to the spoilage during storage.
This study concerns an attempt to establish a new method for predicting antimicrobial peptides (AMPs) which are important to the immune system. Recently, researchers are interested in designing alternative drugs based on AMPs because they have found that a large number of bacterial strains have become resistant to available antibiotics. However, researchers have encountered obstacles in the AMPs designing process as experiments to extract AMPs from protein sequences are costly and require a long set-up time. Therefore, a computational tool for AMPs prediction is needed to resolve this problem. In this study, an integrated algorithm is newly introduced to predict AMPs by integrating sequence alignment and support vector machine- (SVM-) LZ complexity pairwise algorithm. It was observed that, when all sequences in the training set are used, the sensitivity of the proposed algorithm is 95.28% in jackknife test and 87.59% in independent test, while the sensitivity obtained for jackknife test and independent test is 88.74% and 78.70%, respectively, when only the sequences that has less than 70% similarity are used. Applying the proposed algorithm may allow researchers to effectively predict AMPs from unknown protein peptide sequences with higher sensitivity.
Rheumatoid arthritis (RA) is an inflammatory autoimmune disease affecting about 0.24 % of the world population. Protein arginine deiminase type 4 (PAD4) is believed to be responsible for the occurrence of RA by catalyzing citrullination of proteins. The citrullinated proteins act as autoantigens by stimulating an immune response. Citrullinated α-enolase has been identified as one of the autoantigens for RA. Hence, α-enolase serves as a suitable template for design of potential peptide inhibitors against PAD4. The binding affinity of α-enolase-derived peptides and PAD4 was virtually determined using PatchDock and HADDOCK docking programs. Synthesis of the designed peptides was performed using a solid phase peptide synthesis method. The inhibitory potential of each peptide was determined experimentally by PAD4 inhibition assay and IC50 measurement. PAD4 assay data show that the N-P2 peptide is the most favourable substrate among all peptides. Further modification of N-P2 by changing the Arg residue to canavanine [P2 (Cav)] rendered it an inhibitor against PAD4 by reducing the PAD4 activity to 35 % with IC50 1.39 mM. We conclude that P2 (Cav) is a potential inhibitor against PAD4 and can serve as a starting point for the development of even more potent inhibitors.
Antimicrobial peptides (AMPs) are expressed in various living organisms as first-line host defenses against potential harmful encounters in their surroundings. AMPs are short polycationic peptides exhibiting various antimicrobial activities. The principal antibacterial activity is attributed to the membrane-lytic mechanism which directly interferes with the integrity of the bacterial cell membrane and cell wall. In addition, a number of AMPs form a transmembrane channel in the membrane by self-aggregation or polymerization, leading to cytoplasm leakage and cell death. However, an increasing body of evidence has demonstrated that AMPs are able to exert intracellular inhibitory activities as the primary or supportive mechanisms to achieve efficient killing. In this review, we focus on the major intracellular targeting activities reported in AMPs, which include nucleic acids and protein biosynthesis and protein-folding, protease, cell division, cell wall biosynthesis, and lipopolysaccharide inhibition. These multifunctional AMPs could serve as the potential lead peptides for the future development of novel antibacterial agents with improved therapeutic profiles.
BACKGROUND: Despite the extensive research carried out to develop natural antifungal preservatives for food applications, there are very limited antifungal agents available to inhibit the growth of spoilage fungi in processed foods. Scope and Approach: Therefore, this review summarizes the discovery and development of antifungal peptides using lactic acid bacteria fermentation to prevent food spoilage by fungi. The focus of this review will be on the identification of antifungal peptides, potential sources, the possible modes of action and properties of peptides considered to inhibit the growth of spoilage fungi. Key Findings and Conclusions: Antifungal peptides generated by certain lactic acid bacteria strains have a high potential for applications in a broad range of foods. The mechanism of peptides antifungal activity is related to their properties such as low molecular weight, concentration and secondary structure. The antifungal peptides were proposed to be used as bio-preservatives to reduce and/or replace chemical preservatives.
Microbial oils are considered as alternative to vegetable oils or animal fats as biodiesel feedstock. Microalgae and oleaginous yeast are the main candidates of microbial oil producers' community. However, biodiesel synthesis from these sources is associated with high cost and process complexity. The traditional transesterification method includes several steps such as biomass drying, cell disruption, oil extraction and solvent recovery. Therefore, direct transesterification or in situ transesterification, which combines all the steps in a single reactor, has been suggested to make the process cost effective. Nevertheless, the process is not applicable for large-scale biodiesel production having some difficulties such as high water content of biomass that makes the reaction rate slower and hurdles of cell disruption makes the efficiency of oil extraction lower. Additionally, it requires high heating energy in the solvent extraction and recovery stage. To resolve these difficulties, this review suggests the application of antimicrobial peptides and high electric fields to foster the microbial cell wall disruption.
Five Pinto bean peptides with α-amylase and angiotensin converting enzyme (ACE) inhibitory activities were successfully identified using the integrated bioinformatics approach. By using PEAKS studio, 511 peptide sequences were first shortlisted based on their de novo sequence property and average local confidence (ALC) yield of ≥60%. Subsequently, only five peptides were found to have high potential (score ≥0.80) for contributing bioactivy. The important sites which were potentially bound by the peptides: (a) Trp58, Trp59, Tyr 62, Asp96, Arg195, Asp197, Glu233, His299, Asp300 and His305 for α-amylase; (b) His353, Ala354, His383, Glu384, His387, Glu411, Lys511, His513, Tyr520 and Tyr523 for ACE had corresponded to the catalytic and substrate binding sites of the two enzymes. A validation assay was then conducted and IC50 values were determined. The range of the values for α-amylase inhibitory activity was 10.03-23.33mM, whereas the values for ACE inhibitory activity were of 1.52-31.88μM.
Edible bird's nest (EBN) is widely consumed as a delicacy and traditional medicine amongst the Chinese. In the present study, for the first time, the antioxidant properties of an EBN pepsin-trypsin hydrolysate of the swiftlet species Aerodramus fuciphagus and its ultrafiltration fractions were investigated. Thirteen peptides with molecular weights between 514.29 and 954.52 Da were identified in the EBN fraction with the use of mass spectrometry. Two novel pentapeptides Pro-Phe-His-Pro-Tyr and Leu-Leu-Gly-Asp-Pro, corresponding to f134-138 and f164-168 of cytochrome b of A. fuciphagus, indicated the highest ORAC values of 14.95 and 14.32 μM of TE μM(-1) peptide, respectively. Both purified peptides showed resistance against simulated gastrointestinal proteases. In addition, both peptides had no in vitro cytotoxicity on human lung MRC-5 cells and prevented human liver carcinoma HepG2 cellular damage caused by hydroxyl radicals. Therefore, it is suggested that EBN protein hydrolysates are a good source of natural antioxidants and could be applied as nutraceutical compounds.