Browse
Submit Data
Databases
API
Help

Dataset Information

18 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

De novo expression and antibacterial potential of four lactoferricin peptides in cell-free protein synthesis system

ABSTRACT: Highlights • Cell-free (in vitro) method is ideal for short bioactive peptides rapid expression.• Camel lactoferricin is the most active among natural lactoferricin peptides.• Consensus lactoferricin is a candidate for further development into therapeutic use. For the first time, we produced four lactoferricin (LFcin) peptides by a cell-free (in vitro) method. These short antimicrobial peptides were expressed in an E. coli cell-free protein synthesis (CFPS) system and the bioactivity of the produced peptides was demonstrated. Additionally, we designed a novel synthetic consensus peptide (ConLFcin). The genes of bovine Lfcin (bLFcin), human Lfcin (hLFcin), camel Lfcin (cLFcin), and ConLFcin were cloned into pET101/D-TOPO vector then peptides were synthesized in vitro by E. coli CFPS system. The antibacterial activity of these synthesized peptides was evaluated against Escherichia coli, Salmonella typhi, Pseudomonas aeruginosa, Staphylococcus aureus, and methicillin-resistant Staphylococcus aureus (MRSA). The four cell-free synthesized peptides showed significant antibacterial potency at minimum inhibitory concentration (MIC) values between 1.25 and 10 ?g/mL. cLFcin and ConLFcin showed higher antibacterial effects than bLFcin and hLFcin. Thus, cell-free expression system is an ideal system for rapid expression of functionally active short bioactive peptides.

SUBMITTER: El-Baky N

PROVIDER: S-EPMC7779732 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

de novo synthesis of a bacterial toxin/antitoxin system.

Project description:The prevalence of toxin/antitoxin (TA) systems in almost all genomes suggests they evolve rapidly. Here we show that an antitoxin from a type V system (GhoS, an endoribonuclease specific for the mRNA of the toxin GhoT) can be converted into a novel toxin (ArT) simply by adding two mutations. In contrast to GhoS, which increases growth, the new toxin ArT decreases growth dramatically in Escherichia coli. Transmission electron microscopy analysis revealed that the nucleoid in ArT-producing cells is concentrated and appears hollow. Whole-transcriptome profiling revealed ArT cleaves 50 additional transcripts, which shows that the endoribonuclease activity of GhoS has been broadened as it was converted to ArT. Furthermore, we evolved an antitoxin for the new toxin ArT from two unrelated antitoxin templates, the protein-based antitoxin MqsA and RNA-based antitoxin ToxI, and showed that the evolved MqsA and ToxI variants are able to counteract the toxicity of ArT. In addition, the de novo TA system was found to increase persistence, a phenotype commonly associated with TA systems. Therefore, toxins and antitoxins from disparate systems can be interconverted.

| S-EPMC4010927 | biostudies-literature

De novo reconstitution of chromatin using wheat germ cell-free protein synthesis.

Project description:DNA is packaged with histones to form chromatin that impinges on all nuclear processes, including transcription, replication and repair, in the eukaryotic nucleus. A complete understanding of these molecular processes requires analysis of chromatin context in vitro. Here, Drosophila four core histones were produced in a native and unmodified form using wheat germ cell-free protein synthesis. In the assembly reaction, four unpurified core histones and three chromatin assembly factors (dNAP-1, dAcf1 and dISWI) were incubated with template DNA. We then assessed stoichiometry with the histones, nucleosome arrays, supercoiling and the ability of the chromatin to serve as a substrate for histone-modifying enzymes. Overall, our method provides a new avenue to produce chromatin that can be useful in a wide range of chromatin research.

| S-EPMC8167859 | biostudies-literature

Novel antibiotic-free plasmid selection system based on complementation of host auxotrophy in the NAD de novo synthesis pathway.

Project description:The use of antibiotic resistance genes in plasmids causes potential biosafety and clinical hazards, such as the possibility of horizontal spread of resistance genes or the rapid emergence of multidrug-resistant pathogens. This paper introduces a novel auxotrophy complementation system that allowed plasmids and host cells to be effectively selected and maintained without the use of antibiotics. An Escherichia coli strain carrying a defect in NAD de novo biosynthesis was constructed by knocking out the chromosomal quinolinic acid phosphoribosyltransferase (QAPRTase) gene. The resistance gene in the plasmids was replaced by the QAPRTase gene of E. coli or the mouse. As a result, only expression of the QAPRTase gene from plasmids can complement and rescue E. coli host cells in minimal medium. This is the first time that a vertebrate gene has been used to construct a nonantibiotic selection system, and it can be widely applied in DNA vaccine and gene therapy. As the QAPRTase gene is ubiquitous in species ranging from bacteria to mammals, the potential environmental biosafety problems caused by horizontal gene transfer can be eliminated.

| S-EPMC2849241 | biostudies-literature

Toward Molecular Cooperation by De Novo Peptides.

Project description:Theoretical models of the chemical origins of life depend on self-replication or autocatalysis, processes that arise from molecular interactions, recruitment, and cooperation. Such models often lack details about the molecules and reactions involved, giving little guidance to those seeking to detect signs of interaction, recruitment, or cooperation in the laboratory. Here, we develop minimal mathematical models of reactions involving specific chemical entities: amino acids and their condensation reactions to form de novo peptides. Reactions between two amino acids form a dipeptide product, which enriches linearly in time; subsequent recruitment of such products to form longer peptides exhibit super-linear growth. Such recruitment can be reciprocated: a peptide contributes to and benefits from the formation of one or more other peptides; in this manner, peptides can cooperate and thereby exhibit autocatalytic or exponential growth. We have started to test these predictions by quantitative analysis of de novo peptide synthesis conducted by wet-dry cycling of a five-amino acid mixture over 21 days. Using high-performance liquid chromatography, we tracked abundance changes for >60 unique peptide species. Some species were highly transient, with the emergence of up to 17 new species and the extinction of nine species between samplings, while other species persisted across many cycles. Of the persisting species, most exhibited super-linear growth, a sign of recruitment anticipated by our models. This work shows how mathematical modeling and quantitative analysis of kinetic data can guide the search for prebiotic chemistries that have the potential to cooperate and replicate.

| S-EPMC8212187 | biostudies-literature

Fluorescent Amino Acid Initiated de novo Cyclic Peptides for the Label-Free Assessment of Cell Permeability*.

Project description:The major obstacle in applying peptides to intracellular targets is their low inherent cell permeability. Standard approaches to attach a fluorophore (e. g. FITC, TAMRA) can change the physicochemical properties of the parent peptide and influence their ability to penetrate and localize in cells. We report a label-free strategy for evaluating the cell permeability of cyclic peptide leads. Fluorescent tryptophan analogues 4-cyanotryptophan (4CNW) and β-(1-azulenyl)-L-alanine (AzAla) were incorporated into in vitro translated macrocyclic peptides by initiator reprogramming. We then demonstrate these efficient blue fluorescent emitters are good tools for monitoring peptide penetration into cells.

| S-EPMC8597039 | biostudies-literature

Dereplication and de novo sequencing of nonribosomal peptides.

Project description:Nonribosomal peptides (NRPs) are of great pharmacological importance, but there is currently no technology for high-throughput NRP 'dereplication' and sequencing. We used multistage mass spectrometry followed by spectral alignment algorithms for sequencing of cyclic NRPs. We also developed an algorithm for comparative NRP dereplication that establishes similarities between newly isolated and previously identified similar but nonidentical NRPs, substantially reducing dereplication efforts.

| S-EPMC2754211 | biostudies-literature

Accurate de novo design of hyperstable constrained peptides.

Project description:Naturally occurring, pharmacologically active peptides constrained with covalent crosslinks generally have shapes that have evolved to fit precisely into binding pockets on their targets. Such peptides can have excellent pharmaceutical properties, combining the stability and tissue penetration of small-molecule drugs with the specificity of much larger protein therapeutics. The ability to design constrained peptides with precisely specified tertiary structures would enable the design of shape-complementary inhibitors of arbitrary targets. Here we describe the development of computational methods for accurate de novo design of conformationally restricted peptides, and the use of these methods to design 18-47 residue, disulfide-crosslinked peptides, a subset of which are heterochiral and/or N-C backbone-cyclized. Both genetically encodable and non-canonical peptides are exceptionally stable to thermal and chemical denaturation, and 12 experimentally determined X-ray and NMR structures are nearly identical to the computational design models. The computational design methods and stable scaffolds presented here provide the basis for development of a new generation of peptide-based drugs.

| S-EPMC5161715 | biostudies-literature

De novo assembled transcriptomes for four Notothenioid species (Antarctic icefish)

Project description: Not available

| S-BSST132 | biostudies-other

De Novo Synthesis of Benzannelated Heterocycles.

Project description:Benzannelated heterocycles such as indoles and indazoles are prominent structural motifs found in natural products, pharmaceuticals and agrochemicals. For their synthesis, chemists traditionally either functionalize commercially available heterocycles or resort to transformations that make use of benzene-derived building blocks. Here, we report a powerful cascade reaction that enables the de novo construction of variously substituted indoles, indazoles, benzofurans and benzothiophenes from readily available bicyclo[3.1.0]hexan-2-ones. The transformation can be conducted under mild, non-anhydrous conditions. For the synthesis of indoles, mechanistic studies revealed that the electrocyclic ring-opening of the bicyclic ring-system and aromatization precedes the 3,3-sigmatropic rearrangement.

| S-EPMC5821223 | biostudies-literature

Enzymatic de novo pyrimidine nucleotide synthesis.

Project description:The use of stable isotope labeling has revolutionized NMR studies of nucleic acids, and there is a need for methods of incorporation of specific isotope labels to facilitate specific NMR experiments and applications. Enzymatic synthesis offers an efficient and flexible means to synthesize nucleoside triphosphates from a variety of commercially available specifically labeled precursors, permitting isotope labeling of RNAs prepared by in vitro transcription. Here, we recapitulate de novo pyrimidine biosynthesis in vitro, using recombinantly expressed enzymes to perform efficient single-pot syntheses of UTP and CTP that bear a variety of stable isotope labeling patterns. Filtered NMR experiments on (13)C, (15)N, (2)H-labeled HIV-2 TAR RNA demonstrate the utility and value of this approach. This flexible enzymatic synthesis will make implementing detailed and informative RNA stable isotope labeling schemes substantially more cost-effective and efficient, providing advanced tools for the study of structure and dynamics of RNA molecules.

| S-EPMC3134529 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data