Antimicrobial Peptide Database Research Articles

Antimicrobial Peptides: An Update on Classifications and Databases.

Antimicrobial peptides (AMPs) are distributed across all kingdoms of life and are an indispensable component of host defenses. They consist of predominantly short cationic peptides with a wide variety of structures and targets. Given the ever-emerging resistance of various pathogens to existing antimicrobial therapies, AMPs have recently attracted extensive interest as potential therapeutic agents. As the discovery of new AMPs has increased, many databases specializing in AMPs have been developed to collect both fundamental and pharmacological information. In this review, we summarize the sources, structures, modes of action, and classifications of AMPs. Additionally, we examine current AMP databases, compare valuable computational tools used to predict antimicrobial activity and mechanisms of action, and highlight new machine learning approaches that can be employed to improve AMP activity to combat global antimicrobial resistance.

Antifungal Activity of ToAP2D Peptide Against Sporothrix globosa

Improving clinical efficacy and reducing treatment time have been the focus of sporotrichosis therapy. Antimicrobial peptides ToAP2A, ToAP2C, and ToAP2D were synthesized on the basis of ToAP2 (AP02759), a peptide derived from the antimicrobial peptide database by the database filtering technology, and their physicochemical characteristics were analyzed. Compared with template peptide ToAP2, the modified peptides had much shorter length, lower molecular weight but significantly greater stability, which in return resulted in increases in the aliphatic index, hydrophilicity, and protein binding ability. Here, we show that the three derived peptides inhibit the growth of Sporothrix globosa, among which ToAP2D had the strongest anti-fungal activity. ToAP2D showed good serum stability without acute toxicity. The ToAP2D treatment inhibited the growth of S. globosa and enhanced apoptosis, which was evidenced by the upregulation of apoptosis-related protein caspase-3. The scanning electron microscopy analysis revealed deformation and rupture of S. globosa. The levels of mitochondrial membrane potential were decreased and that of the reactive oxygen species (ROS) were increased in S. globosa upon ToAP2D treatment. Moreover, ToAP2D activated metacaspase. In the in vivo study, we further demonstrated that ToAP2D inhibited the S. globosa infection of mice footpads, and its efficiency was nearly comparable to itraconazole. In summary, our results suggest that antimicrobial peptide ToAP2D has the potential for sporotrichosis therapy.

Identification of Antimicrobial Peptide Genes in Black Rockfish Sebastes schlegelii and Their Responsive Mechanisms to Edwardsiella tarda Infection.

Simple SummarySebastes schlegelii is a typical viviparous teleost that belongs to the family Scorpaenidae, and is an important mariculture fish in East Asian countries such as Japan, Korea, and China because of its high economic and ecological values. With the enlargement of cultivation scale, bacterial and viral diseases have become the main threats to the farming industry of S. schlegelii, which resulted in significant economic losses. The genome features and evolutionary position of S. schlegelii were investigated based on multiple sequencing approaches. Then, genome-wide identification of AMPs was performed according to the known AMPs in the Antimicrobial Peptides Database. Furthermore, the expression levels of these AMPs were detected to explore their responsive mechanisms to pathogenic bacteria. The availability of the high-quality genome of S. schlegelii will be valuable for understanding its genome features and evolutionary history. Genome-wide AMP identification could be beneficial for understanding the molecular mechanism of innate and adaptive immune responses of S. schlegelii.The black rockfish, Sebastes schlegelii, is a typical viviparous teleost, which belongs to the family Scorpaenidae. Due to its high economic and ecological values, S. schlegelii has been widely cultured in East Asian countries. With the enlargement of cultivation scale, bacterial and viral diseases have become the main threats to the farming industry of S. schlegelii, which have resulted in significant economic losses. In this study, Illumina shotgun sequencing, single-molecule real-time (SMRT) sequencing, 10× genomics and high-throughput chromosome conformation capture (Hi-C) technologies were collectively applied to assemble the genome of S. schlegelii. Then, we identified the antimicrobial peptide genes (AMPs) in the S. schlegelii genome. In total, 214 AMPs were identified in the S. schlegelii genome, which can be divided into 33 classes according to the annotation and cataloging of the Antimicrobial Peptides Database (APD3). Among these AMPs, thrombin-derived C-terminal peptide (TCP) was the dominant type, followed by RegIIIgamma and chemokine. The amino acid sequences of the TCP, cgUbiquitin, RegIIIalpha, RegIIIgamma, chemokine shared 32.55%, 42.63%, 29.87%, 28.09%, and 32.15% similarities among the same type in S. schlegelii. Meanwhile, the expression patterns of these AMPs in nine healthy tissues and at different infection time points in intestine were investigated. The results showed that the numbers and types of AMPs that responded to Edwardsiella tarda infection gradually increased as the infection progressed. In addition, we analyzed the phylogenetic relationships of hepcidins in teleost. The identification of AMPs based on the whole genome could provide a comprehensive database of potential AMPs, and benefit for the understanding of the molecular mechanisms of immune responses to E. tarda infection in S. schlegelii. This would further offer insights into an accurate and effective design and development of AMP for aquaculture therapy in the future.

The evolution of the antimicrobial peptide database over 18 years: Milestones and new features.

The antimicrobial peptide database (APD) has served the antimicrobial peptide field for 18 years. Because it is widely used in research and education, this article documents database milestones and key events that have transformed it into the current form. A comparison is made for the APD peptide statistics between 2010 and 2020, validating the major database findings to date. We also describe new additions ranging from peptide entries to search functions. Of note, the APD also contains antimicrobial peptides from host microbiota, which are important in shaping immune systems and could be linked to a variety of human diseases. Finally, the database has been re-programmed to the web branding and latest security compliance of the University of Nebraska Medical Center. The reprogrammed APD can be accessed at https://aps.unmc.edu.

DRAMP 3.0: an enhanced comprehensive data repository of antimicrobial peptides.

Stapled antimicrobial peptides are an emerging class of artificial cyclic peptide molecules which have antimicrobial activity and potent structure stability. We previously published the Data Repository of Antimicrobial Peptides (DRAMP) as a manually annotated and open-access database of antimicrobial peptides (AMPs). In the update of version 3.0, special emphasis was placed on the new development of stapled AMPs, and a subclass of specific AMPs was added to store information on these special chemically modified AMPs. To help design low toxicity AMPs, we also added the cytotoxicity property of AMPs, as well as the expansion of newly discovered AMP data. At present, DRAMP has been expanded and contains 22259 entries (2360 newly added), consisting of 5891 general entries, 16110 patent entries, 77 clinical entries and 181 stapled AMPs. A total of 263 entries have predicted structures, and more than 300 general entries have links to experimentally determined structures in the Protein Data Bank. The update also covers new annotations, statistics, categories, functions and download links. DRAMP is available online at http://dramp.cpu-bioinfor.org/.

Nanostructured Antimicrobial Peptides: Crucial Steps of Overcoming the Bottleneck for Clinics.

The security issue of human health is faced with dispiriting threats from multidrug-resistant bacteria infections induced by the abuse and misuse of antibiotics. Over decades, the antimicrobial peptides (AMPs) hold great promise as a viable alternative to treatment with antibiotics due to their peculiar antimicrobial mechanisms of action, broad-spectrum antimicrobial activity, lower drug residue, and ease of synthesis and modification. However, they universally express a series of disadvantages that hinder their potential application in the biomedical field (e.g., low bioavailability, poor protease resistance, and high cytotoxicity) and extremely waste the abundant resources of AMP database discovered over the decades. For all these reasons, the nanostructured antimicrobial peptides (Ns-AMPs), based on a variety of nanosystem modification, have made up for the deficiencies and pushed the development of novel AMP-based antimicrobial therapies. In this review, we provide an overview of the advantages of Ns-AMPs in improving therapeutic efficacy and biological stability, reducing side effects, and gaining the effect of organic targeting and drug controlled release. Then the different material categories of Ns-AMPs are described, including inorganic material nanosystems containing AMPs, organic material nanosystems containing AMPs, and self-assembled AMPs. Additionally, this review focuses on the Ns-AMPs for the effect of biological activities, with emphasis on antimicrobial activity, biosecurity, and biological stability. The “state-of-the-art” antimicrobial modes of Ns-AMPs, including controlled release of AMPs under a specific environment or intrinsic antimicrobial properties of Ns-AMPs, are also explicated. Finally, the perspectives and conclusions of the current research in this field are also summarized.

In-silico study of peptide-protein interaction of antimicrobial peptides potentially targeting SARS and SARS-CoV-2 nucleocapsid protein.

This study is an attempt to find a suitable therapy using antimicrobial peptides (AMPs) by identifying peptide-protein interaction of AMPs and nucleocapsid protein of SARS and SARS-CoV- 2. The AMPs were shortlisted from the APD3 database (Antimicrobial peptide database) based on various physicochemical parameters. The binding efficacy of AMPs was measured using the lowest energy score of the docked complexes with 10 selected AMPs. For SARS-CoV, AP00180 showed the best pose with a binding affinity value of − 6.4 kcal/mol. Prominent hydrogen bonding interactions were observed between Lys85 (nucleocapsid receptor) and Arg13 (antimicrobial peptide ligand) having the least intermolecular distance of 1.759 Å. For SARS-CoV-2, AP00549 was docked with a binding affinity value of − 3.4 kcal/mol and Arg119 and Glu14 of receptor nucleocapsid protein and ligand AMP having the least intermolecular distance of 2.104 The dynamic simulation was performed at 50 ns to check the stability of the final docked complexes, one with each protein. The two best AMPs were AP00180 (Human Defensin-5) for SARS and AP00549 (Plectasin) for SARS-CoV-2. From positive results of dynamic simulation and previously known knowledge that some AMPs interact with the nucleocapsid of coronaviruses, these AMPs might be used as a potential therapeutic agent for the treatment regime of SARS-CoV-2 and SARS infection.Supplementary InformationThe online version contains supplementary material available at 10.1007/s40203-021-00103-z.

Computational Methods and Tools in Antimicrobial Peptide Research.

The evolution of antibiotic-resistant bacteria is an ongoing and troubling development that has increased the number of diseases and infections that risk going untreated. There is an urgent need to develop alternative strategies and treatments to address this issue. One class of molecules that is attracting significant interest is that of antimicrobial peptides (AMPs). Their design and development has been aided considerably by the applications of molecular models, and we review these here. These methods include the use of tools to explore the relationships between their structures, dynamics, and functions and the increasing application of machine learning and molecular dynamics simulations. This review compiles resources such as AMP databases, AMP-related web servers, and commonly used techniques, together aimed at aiding researchers in the area toward complementing experimental studies with computational approaches.

A Novel Antimicrobial Peptides From Pine Needles of Pinus densiflora Sieb. et Zucc. Against Foodborne Bacteria.

Pine needles are used in several East Asian countries as food or traditional medicine. It contains functional components that exhibit a wide spectrum of pharmacological effects such as antioxidant, antimicrobial, anti-diabetic, and anti-inflammatory activities. We determined and characterized the novel antimicrobial peptides (AMPs) isolated from Pinus densiflora Sieb. et Zucc. The four active pine-needle (PN) peptides showed antimicrobial activity against foodborne bacteria with minimum inhibitory concentration (MIC) values within the range of 8–128 μg/ml. PN peptides showed no detectable hemolytic activity or cytotoxicity at the antimicrobial concentrations. The N-terminal amino acid sequence of the PN5 was identified using Edman degradation and Antimicrobial Peptide Database (APD) homology analysis showed that it was not identical to any other plant peptide. This suggests that PN5 can serve as an alternative therapeutic agent to be used in the food industry.

Synthetic short peptides (SSPs) as antibiofilm agents for dental material applications

Human oral microflora is one of the most complex microbial communities known. A consistent competitive environment in this niche necessitates the generation of bacteriocins or antimicrobial peptides (AMPs), which play a significant impact on niche ecology by inhibiting the growth of closely related inhabitants. In the current study, draft genomes of oral commensal Streptococcus salivarius strains were analyzed using various bioinformatic tools to design novel short antibiofilm peptides. The antimicrobial potency of peptides was predicted by using AMP database and Bactibase. Novel short peptides were designed, synthesized and evaluated against oral oppurtinistic pathogenic microbes. Synthetic short peptides (SSPs) displayed effective inhibition of biofilm formation by specific oral bacterial strains, indicating their potential applicability as targeted antibiofilm agents for variety of dental material surface coating applications.

Exploring effectiveness of ab-initio protein-protein docking methods on a novel antibacterial protein complex dataset.

Diseases caused by bacterial infections become a critical problem in public heath. Antibiotic, the traditional treatment, gradually loses their effectiveness due to the resistance. Meanwhile, antibacterial proteins attract more attention because of broad spectrum and little harm to host cells. Therefore, exploring new effective antibacterial proteins is urgent and necessary. In this paper, we are committed to evaluating the effectiveness of ab-initio docking methods in antibacterial protein-protein docking. For this purpose, we constructed a three-dimensional (3D) structure dataset of antibacterial protein complex, called APCset, which contained $19$ protein complexes whose receptors or ligands are homologous to antibacterial peptides from Antimicrobial Peptide Database. Then we selected five representative ab-initio protein-protein docking tools including ZDOCK3.0.2, FRODOCK3.0, ATTRACT, PatchDock and Rosetta to identify these complexes' structure, whose performance differences were obtained by analyzing from five aspects, including top/best pose, first hit, success rate, average hit count and running time. Finally, according to different requirements, we assessed and recommended relatively efficient protein-protein docking tools. In terms of computational efficiency and performance, ZDOCK was more suitable as preferred computational tool, with average running time of $6.144$ minutes, average Fnat of best pose of $0.953$ and average rank of best pose of $4.158$. Meanwhile, ZDOCK still yielded better performance on Benchmark 5.0, which proved ZDOCK was effective in performing docking on large-scale dataset. Our survey can offer insights into the research on the treatment of bacterial infections by utilizing the appropriate docking methods.

Construction of Unified Human Antimicrobial and Immunomodulatory Peptide Database and Examination of Antimicrobial and Immunomodulatory Peptides in Alzheimer's Disease Using Network Analysis of Proteomics Datasets.

The reanalysis of genomics and proteomics datasets by bioinformatics approaches is an appealing way to examine large amounts of reliable data. This can be especially true in cases such as Alzheimer’s disease, where the access to biological samples, along with well-defined patient information can be challenging. Considering the inflammatory part of Alzheimer’s disease, our aim was to examine the presence of antimicrobial and immunomodulatory peptides in human proteomic datasets deposited in the publicly available proteomics database ProteomeXchange (http://www.proteomexchange.org/). First, a unified, comprehensive human antimicrobial and immunomodulatory peptide database, containing all known human antimicrobial and immunomodulatory peptides was constructed and used along with the datasets containing high-quality proteomics data originating from the examination of Alzheimer’s disease and control groups. A throughout network analysis was carried out, and the enriched GO functions were examined. Less than 1% of all identified proteins in the brain were antimicrobial and immunomodulatory peptides, but the alterations characteristic of Alzheimer’s disease could be recapitulated with their analysis. Our data emphasize the key role of the innate immune system and blood clotting in the development of Alzheimer’s disease. The central role of antimicrobial and immunomodulatory peptides suggests their utilization as potential targets for mechanistic studies and future therapies.

Novel Chronic Wound Healing by Anti-biofilm Peptides and Protease.

Chronic wounds have made a challenge in medical healthcare due to their biofilm infections, which reduce the penetrance of the antibacterial agents in the injury site. In infected wounds, the most common bacterial strains are Staphylococcus aureus and Pseudomonas aeruginosa. Biofilm disruption in chronic wounds is crucial in wound healing. Due to their broad-spectrum antibacterial properties and fewer side effects, anti-biofilm peptides, especially bacteriocins, are promising in the healing of chronic wounds by biofilm destruction. This study reviews the effects of antimicrobial and anti-biofilm agents, including bacteriocins and protease enzymes as a novel approach, on wound healing, along with analyzing the molecular docking between a bacterial protease and biofilm components. Among a large number of anti-biofilm bacteriocins identified up to now, seven types have been registered in the antimicrobial peptides (AMPs) database. Although it is believed that bacterial proteases are harmful in wound healing, it has recently been demonstrated that these proteases like the human serine protease, in combination with AMPs, can improve wound healing by biofilm destruction. In this work, docking results between metalloprotease from Paenibacillus polymyxa and proteins of S. aureus and P. aeruginosa involved in biofilm production, showed that this bacterial protease could efficiently interact with biofilm components. Infected wound healing is an important challenge in clinical trials due to biofilm production by bacterial pathogens. Therefore, simultaneous use of proteases or anti-biofilm peptides with antimicrobial agents could be a promising method for chronic wound healing.

Broad-Spectrum Robust Direct Bactericidal Activity of Fish IFNφ1 Reveals an Antimicrobial Peptide-like Function for Type I IFNs in Vertebrates.

Type I IFNs (IFN-Is) play pivotal roles in host defense against viral infections but remain enigmatic against bacterial pathogens. In this study, we recombinantly expressed and purified intact grass carp (Ctenopharyngodon idella) IFNφ1 (gcIFNφ1), a teleost IFN-I. gcIFNφ1 widely powerfully directly kills both Gram-negative and Gram-positive bacteria in a dose-dependent manner. gcIFNφ1 binds to LPS or peptidoglycan and provokes bacterial membrane depolarization and disruption, resulting in bacterial death. Furthermore, gcIFNφ1 can efficiently protect zebrafish against Aeromonas hydrophila infection and significantly reduce the bacterial loads in tissues by an infection model. In addition, we wonder whether antibacterial IFN-I members exist in other vertebrates. The amino acid compositions of representative IFN-Is with strong positive charges from Pisces, Amphibia, reptiles, Aves, and Mammalia demonstrate high similarities with those of 2237 reported cationic antimicrobial peptides in antimicrobial peptide database. Recombinant intact representative IFN-I members from the nonmammalian sect exhibit potent broad-spectrum robust bactericidal activity through bacterial membrane depolarization; in contrast, the bactericidal activity is very weak from mammalian IFN-Is. The findings display a broad-spectrum potent direct antimicrobial function for IFN-Is, to our knowledge previously unknown. The results highlight that IFN-Is are important and robust in host defense against bacterial pathogens, and unify direct antibacterial and indirect antiviral bifunction in nonmammalian jawed vertebrates.

In silico strategic curation, retrieval and prediction of novel antimicrobial peptide from Locusta migratoria transcriptome

Abstract Evolutionary immune system upgradation is done by all living organism to survive from the invading pathogens. Insects has a strong defense system comprising of anatomical barrier, humoral antimicrobial peptide (AMP) production and cellular immune components. Locusta migratoria (L. migratoria) is an agricultural pest insect migrating long distance might encounter distinct pathogens. Owing to this, we aimed in identifying AMPs present in the L. migratoria and elucidate the antimicrobial activity. The migratory locust was immunized with Escherichia coli, Staphylococcus aureus, Candida albicans mixture, then RNA was isolated and sequenced. The L. migratoria transcriptome was de novo assembled using trinity and screened for AMPs propensity specifically molecular and aggregation properties. The novelty is tested by blasting in known AMP databases. Finally, obtained novel putative AMPs were then tested for antimicrobial and hemolytic activity. The prediction resulted in 3,524 putative AMPs which was further screened down to ten AMPs for final testing. Two peptides showing promising antimicrobial effects were obtained. All the analyzed peptides showed no hemolysis confirms its non-toxicity. Thus, our peptides could be promising drug candidate and can be used as an alternative to antibacterial or antifungal therapy with further validations.

New Putative Antimicrobial Candidates: In silico Design of Fish-Derived Antibacterial Peptide-Motifs.

Antimicrobial resistance remains a great threat to global health. In response to the World Health Organizations’ global call for action, nature has been explored for novel and safe antimicrobial candidates. To date, fish have gained recognition as potential source of safe, broad spectrum and effective antimicrobial therapeutics. The use of computational methods to design antimicrobial candidates of industrial application has however, been lagging behind. To fill the gap and contribute to the current fish-derived antimicrobial peptide repertoire, this study used Support Vector Machines algorithm to fish out fish-antimicrobial peptide-motif candidates encrypted in 127 peptides submitted at the Antimicrobial Peptide Database (APD3), steered by their physico-chemical characteristics (i.e., positive net charge, hydrophobicity, stability, molecular weight and sequence length). The best two novel antimicrobial peptide-motifs (A15_B, A15_E) with the lowest instability index (−28.25, −22.49, respectively) and highest isoelectric point (pI) index (10.48 for each) were selected for further analysis. Their 3D structures were predicted using I-TASSER and PEP-FOLD servers while ProSA, PROCHECK, and ANOLEA were used to validate them. The models predicted by I-TASSER were found to be better than those predicted by PEP-FOLD upon validation. Two I-TASSER models with the lowest c-score of −0.10 and −0.30 for A15_B and A15_E peptide-motifs, respectively, were selected for docking against known bacterial-antimicrobial target-proteins retrieved from protein databank (PDB). Carbapenam-3-carboxylate synthase (PDB ID; 4oj8) yielded the lowest docking energy (−8.80 and −7.80 Kcal/mol) against motif A15_B and A15_E, respectively, using AutoDock VINA. Further, in addition to Carbapenam-3-carboxylate synthase, these peptides (A15_B and A15_E) were found to as well bind to membrane protein (PDB ID: 1by3) and Carbapenem synthetase (PDB: 1q15) when ClusPro and HPEPDOCK tools were used. The membrane protein yielded docking energy scores (DES): −290.094, −270.751; coefficient weight (CW): −763.6, 763.3 for A15_B and A15_E) whereas, Carbapenem synthetase (PDB: 1q15) had a DES of −236.802, −262.75 and a CW of −819.7, −829.7 for peptides A15_B and A15_E, respectively. Motif A15_B of amino acid positions 2–19 in Pleurocidin exhibited the strongest in silico antimicrobial potentials. This segment could be a good biological candidate of great application in pharmaceutical industries as an antimicrobial drug candidate.

Data Employed in the Construction of a Composite Protein Database for Proteogenomic Analyses of Cephalopods Salivary Apparatus

Here we provide all datasets and details applied in the construction of a composite protein database required for the proteogenomic analyses of the article “Putative Antimicrobial Peptides of the Posterior Salivary Glands from the Cephalopod Octopus vulgaris Revealed by Exploring a Composite Protein Database”. All data, subdivided into six datasets, are deposited at the Mendeley Data repository as follows. Dataset_1 provides our composite database “All_Databases_5950827_sequences.fasta” derived from six smaller databases composed of (i) protein sequences retrieved from public databases related to cephalopods’ salivary glands, (ii) proteins identified with Proteome Discoverer software using our original data obtained by shotgun proteomic analyses of posterior salivary glands (PSGs) from three Octopus vulgaris specimens (provided as Dataset_2) and (iii) a non-redundant antimicrobial peptide (AMP) database. Dataset_3 includes the transcripts obtained by de novo assembly of 16 transcriptomes from cephalopods’ PSGs using CLC Genomics Workbench. Dataset_4 provides the proteins predicted by the TransDecoder tool from the de novo assembly of 16 transcriptomes of cephalopods’ PSGs. Further details about database construction, as well as the scripts and command lines used to construct them, are deposited within Dataset_5 and Dataset_6. The data provided in this article will assist in unravelling the role of cephalopods’ PSGs in feeding strategies, toxins and AMP production.

In Silico Discovery of Antimicrobial Peptides as an Alternative to Control SARS-CoV-2.

A serious pandemic has been caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The interaction between spike surface viral protein (Sgp) and the angiotensin-converting enzyme 2 (ACE2) cellular receptor is essential to understand the SARS-CoV-2 infectivity and pathogenicity. Currently, no drugs are available to treat the infection caused by this coronavirus and the use of antimicrobial peptides (AMPs) may be a promising alternative therapeutic strategy to control SARS-CoV-2. In this study, we investigated the in silico interaction of AMPs with viral structural proteins and host cell receptors. We screened the antimicrobial peptide database (APD3) and selected 15 peptides based on their physicochemical and antiviral properties. The interactions of AMPs with Sgp and ACE2 were performed by docking analysis. The results revealed that two amphibian AMPs, caerin 1.6 and caerin 1.10, had the highest affinity for Sgp proteins while interaction with the ACE2 receptor was reduced. The effective AMPs interacted particularly with Arg995 located in the S2 subunits of Sgp, which is key subunit that plays an essential role in viral fusion and entry into the host cell through ACE2. Given these computational findings, new potentially effective AMPs with antiviral properties for SARS-CoV-2 were identified, but they need experimental validation for their therapeutic effectiveness.

Design of bovine lactoferricin-derived peptide and its expression and activity in Pichia pastoris

Bovine lactoferrin peptide has been shown to be a broad-spectrum antimicrobial peptide. Based on the relationship between the structure and function of antimicrobial peptides, the antimicrobial peptide databases and protein analysis software were used to optimize the design of bovine lactoferricin peptide (LfcinB). The designed bovine lactoferricin-derived peptide (LfcinBD) gene fragment was inserted into a pPIC9K-His plasmid to construct a recombinant expression vector. After linearization of the Recombinant plasmid, Pichia pastoris GS115 cells were transfected with linearized recombinant plasmid by using electroporation and LfcinBD gene expression was induced with methanol. After the fermentation, supernatant was separated by low-temperature high-speed centrifugation. Ultrafiltration and freeze drying of the fermentation supernatant were performed, purified. Experimental results showed that the LfcinBD had stronger bacteriostatic activity against Staphylococcus aureus than the natural bovine lactoferrin peptide (LfcinB) produced under the same fermentation conditions. The effective expression of the optimized bovine lactoferricin-derived peptide was detected using SDS-PAGE electrophoresis. This study lays the foundation for further exploration to improve the biological activities of antimicrobial peptides.

Putative Antimicrobial Peptides of the Posterior Salivary Glands from the Cephalopod Octopus vulgaris Revealed by Exploring a Composite Protein Database.

Cephalopods, successful predators, can use a mixture of substances to subdue their prey, becoming interesting sources of bioactive compounds. In addition to neurotoxins and enzymes, the presence of antimicrobial compounds has been reported. Recently, the transcriptome and the whole proteome of the Octopus vulgaris salivary apparatus were released, but the role of some compounds—e.g., histones, antimicrobial peptides (AMPs), and toxins—remains unclear. Herein, we profiled the proteome of the posterior salivary glands (PSGs) of O. vulgaris using two sample preparation protocols combined with a shotgun-proteomics approach. Protein identification was performed against a composite database comprising data from the UniProtKB, all transcriptomes available from the cephalopods’ PSGs, and a comprehensive non-redundant AMPs database. Out of the 10,075 proteins clustered in 1868 protein groups, 90 clusters corresponded to venom protein toxin families. Additionally, we detected putative AMPs clustered with histones previously found as abundant proteins in the saliva of O. vulgaris. Some of these histones, such as H2A and H2B, are involved in systemic inflammatory responses and their antimicrobial effects have been demonstrated. These results not only confirm the production of enzymes and toxins by the O. vulgaris PSGs but also suggest their involvement in the first line of defense against microbes.