Identification of cancer-specific motifs in mimotope profiles of serum antibody repertoire.

BACKGROUND: For fighting cancer, earlier detection is crucial. Circulating auto-antibodies produced by the patient's own immune system after exposure to cancer proteins are promising bio-markers for the early detection of cancer. Since an antibody recognizes not the whole antigen but 4-7 critical amino acids within the antigenic determinant (epitope), the whole proteome can be represented by a random peptide phage display library. This opens the possibility to develop an early cancer detection test based on a set of peptide sequences identified by comparing cancer patients' and healthy donors' global peptide profiles of antibody specificities. RESULTS: Due to the enormously large number of peptide sequences contained in global peptide profiles generated by next generation sequencing, the large number of cancer and control sera is required to identify cancer-specific peptides with high degree of statistical significance. To decrease the number of peptides in profiles generated by nextgen sequencing without losing cancer-specific sequences we used for generation of profiles the phage library enriched by panning on the pool of cancer sera. To further decrease the complexity of profiles we used computational methods for transforming a list of peptides constituting the mimotope profiles to the list motifs formed by similar peptide sequences. CONCLUSION: We have shown that the amino-acid order is meaningful in mimotope motifs since they contain significantly more peptides than motifs among peptides where amino-acids are randomly permuted. Also the single sample motifs significantly differ from motifs in peptides drawn from multiple samples. Finally, multiple cancer-specific motifs have been identified.

Development of advanced cardiac progenitor cell culture system through fibronectin and vitronectin derived peptide coated plate.

Cardiovascular disease remains a global health concern. Stem cell therapy utilizing human cardiac progenitor cells (hCPCs) shows promise in treating cardiac vascular disease. However, limited availability and senescence of hCPCs hinder their widespread use. To address these challenges, researchers are exploring innovative approaches. In this study, a bioengineered cell culture plate was developed to mimic the natural cardiac tissue microenvironment. It was coated with a combination of extracellular matrix (ECM) peptide motifs and mussel adhesive protein (MAP). The selected ECM peptide motifs, derived from fibronectin and vitronectin, play crucial roles in hCPCs. Results revealed that the Fibro-P and Vitro-P coated plates significantly improved hCPC adhesion, proliferation, migration, and differentiation compared to uncoated plates. Additionally, long-term culture on the coated plates delayed cellular senescence and maintained hCPC stemness. These enhancements were attributed to the activation of integrin downstream signaling pathways. The findings suggest that the engineered ECM peptide motif-MAP-coated plates hold potential for enhancing the therapeutic efficacy of stem cell-based therapies in cardiac tissue engineering and regenerative medicine.

Identification of two short peptide motifs from serine/arginine-rich protein ribonucleic acid recognition motif-1 domain acting as splicing regulators.

Background: Serine/arginine-rich (SR) proteins regulate pre-mRNA splicing. However, structurally similar proteins often behave differently in splicing regulation and the underlying mechanisms are largely unknown. Here, using SMN1/2 minigenes we extensively analyzed four SR proteins, SRSF1/5/6/9. Methods: In this study, the effects of these proteins on SMN1/2 exon 7 splicing when tethered at either intron 6 or 7 were evaluated using an MS2-tethering assay. Deletion analysis in four SR proteins and co-overexpression analysis were performed. Results: Splicing outcomes varied among all four SR proteins, SRSF1 and SRSF5 function the same at the two sites, acting as repressor and stimulator, respectively; while SRSF6 and SRSF9 promote exon 7 inclusion at only one site. Further, the key domains of each SR proteins were investigated, which identified a potent inhibitory nonapeptide in the C-terminus of SRSF1/9 ribonucleic acid recognition motif-1 (RRM1) and a potent stimulatory heptapeptide at the N-terminus of SRSF5/6 RRM1. Conclusion: The insight of the four SR proteins and their domains in affecting SMN gene splicing brings a new perspective on the modes of action of SR proteins; and the functional peptides obtained here offers new ideas for developing splice switching-related therapies.

Computational Modeling as a Tool to Investigate PPI: From Drug Design to Tissue Engineering.

Protein-protein interactions (PPIs) mediate a large number of important regulatory pathways. Their modulation represents an important strategy for discovering novel therapeutic agents. However, the features of PPI binding surfaces make the use of structure-based drug discovery methods very challenging. Among the diverse approaches used in the literature to tackle the problem, linear peptides have demonstrated to be a suitable methodology to discover PPI disruptors. Unfortunately, the poor pharmacokinetic properties of linear peptides prevent their direct use as drugs. However, they can be used as models to design enzyme resistant analogs including, cyclic peptides, peptide surrogates or peptidomimetics. Small molecules have a narrower set of targets they can bind to, but the screening technology based on virtual docking is robust and well tested, adding to the computational tools used to disrupt PPI. We review computational approaches used to understand and modulate PPI and highlight applications in a few case studies involved in physiological processes such as cell growth, apoptosis and intercellular communication.

In Situ functionalization of Poly(hydroxyethyl methacrylate) Cryogels with Oligopeptides via ß-Cyclodextrin-Adamantane Complexation for Studying Cell-Instructive Peptide Environment.

Oligopeptides are versatile cell modulators resembling pleiotropic activities of ECM proteins and growth factors. Studying the role of cell-instructive peptide signals within 3D scaffolds, yet poorly known, requires effective approaches to introducing bioactive sequences into appropriate materials. We synthesized RGD and GHK motif based peptides 1 and 2 linked to the terminal adamantyl group (Ad) and their fluorescent derivatives 3 and 4. Poly(hydroxyethyl methacrylate) (pHEMA) cryogels with additional PEG/ß-cyclodextrin (CD) units were prepared as an inert macroporous scaffold capable to bind the adamantylated peptides via affinity CD-Ad complexation. According to toluidine blue staining, the CD moieties were effectively and stably incorporated in the pHEMA cryogels at nanomolar amounts per milligram of material. The CD component gradually increased the thickness and swelling ability of the polymer walls of cryogels, resulting in a noticeable decrease in macropore size and modulation of viscoelastic properties. The labeled peptides exhibited fast kinetics of specific binding to the CD-modified cryogels and were simultaneously immobilized by coincubation. The peptide loading approached ca. 0.31 mg per cm2 of cryogel sheet. A well-defined mitogenic effect of the immobilized peptides (2 < 1≪ 1 + 2) was revealed toward 3T3 and PC-12 cells. The synergistic action of RGD and GHK peptides induced a profound change in cell behavior/morphology attributed to a growth-factor-like activity of the composition. Altogether, our results provide an effective procedure for the preparation of CD-modified pHEMA cryogels and their uniform in situ functionalization with bioactive peptide(s) of interest and an informative study of cellular responses in the functionalized scaffolds.

Identification of Ixodid Tick-Specific Aquaporin-1 Potential Anti-tick Vaccine Epitopes: An in-silico Analysis.

Ticks are arthropod vectors of pathogens of both Veterinary and Public health importance. Acaricide application, which is currently the mainstay of tick control, is hampered by high cost, the need for regular application and a selection of multi-acaricide resistant tick populations. In light of this, future tick control approaches are poised to rely on the integration of rational acaricide application and other methods, such as vaccination. To contribute to systematic research-guided efforts to produce anti-tick vaccines, we carried out an in-silico analysis of tick aquaporin-1 (AQP1) protein in order to identify tick-specific AQP1 peptide motifs that can be used in future peptide anti-tick vaccine development. We carried out multiple sequence alignment (MSA), motif analysis, homology modeling, and structural analysis to identify tick-specific AQP1 peptide motifs. BepiPred, Chou and Fasman-Turn, Karplus and Schulz Flexibility, and Parker-Hydrophilicity prediction models were used to predict these motifs' potential to induce B cell mediated immune responses. The tick AQP1 (GenBankID: QDO67142.1) protein was largely similar to the bovine AQP1 (PDB:1J4N) (23 % sequence similarity; Structural superimposition of the homology model and 14JN homotetramers gave an RMSD = 3.269 while superimposition of a single chain gave an RMSD = 1.475). Tick and bovine AQP1 transmembrane domains were largely similar while their extracellular and cytoplasmic domain loops showed variation. Two tick-specific AQP1 peptide motifs; M7 (residues 106-125, p = 5.4e-25), and M8 (residues 85-104, p = 3.3e-24) were identified. These two motifs are located on the extra-cellular AQP1 domain. Motifs; M7 and M8 showed the highest Parker-Hydrophilicity prediction immunogenicity scores of 1.784 and 1.536, respectively. These two motifs can be a good starting point for the development of potential tick AQP1 peptide-based anti-tick vaccines. Further analyses such as molecular dynamics, in vitro assays, and in vivo immunization assays are required to validate these findings.

Screening and Identification of PLK1-Polo Box Binding Peptides by High-Throughput Sequencing of Phage-Selected Libraries.

BACKGROUND: Human proteome contains a plethora of short linear peptide motifs that is crucial for signaling and other cellular processes. These motifs are difficult to identify due to lack of systematic approach for their detection. OBJECTIVES: Here we demonstrate the use of peptide phage display in combination with high throughput next generation sequencing to identify enriched peptide sequences through biopanning process against polo box domain (PBD) of mitotic polo like kinase 1 (Plk1). METHODS: Purified recombinant Plk1 and two unrelated controls namely B-lymphocyte antigen (CD20) and fluorescent protein (mCherry) were subjected to peptide phage display analysis. Bacterially-propagated phage DNA was amplified by PCR using triplet bar coded primers to tag the pool from each amplicon. RESULTS: Proteomic peptide phage display along with next generation sequencing and Bioinformatics analysis demonstrated several known and putative novel interactions which were potentially related to Plk1-PBD. With our strategy, we were able to identify and characterize several Plk1-PBD binding peptides, as well as define more precisely, consensus sequences. CONCLUSION: We believe that this information could provide valuable tools for exploring novel interaction involved in Plk1 signaling as well as to choose peptides for Plk1 specific drug development.

Directed Self-Assembly of Trimeric DNA-Bindingchiral Miniprotein Helicates.

We propose that peptides are highly versatile platforms for the precise design of supramolecular metal architectures, and particularly, for the controlled assembly of helicates. In this context, we show that the bacteriophage T4 Fibritin foldon (T4Ff) can been engineered on its N-terminus with metal-chelating 2,2'-bipyridine units that stereoselectively assemble in the presence of Fe(II) into parallel, three-stranded peptide helicates with preferred helical orientation. Modeling studies support the proposed self-assembly and the stability of the final helicate. Furthermore, we show that these designed mini-metalloproteins selectively recognize three-way DNA junctions over double-stranded DNA.

Modes of MAPK substrate recognition and control.

Mitogen-activated protein kinase (MAPK) cascades are universal, evolutionary conserved signalling modules, which translate environmental information into appropriate responses via phosphorylation of their substrate proteins. In Arabidopsis, the MAPK MPK3 regulates numerous cellular processes, including the adaptation to abiotic and biotic stresses. The molecular steps immediately downstream of MPK3 induction have, therefore, received abundant attention, and a respectable number of MPK3 targets are known by now. These proteins illustrate the substrate promiscuity of MPK3. They also are evidence for how manifold phosphorylation-regulated functions can be. This review presents the current knowledge about the function and regulation of MPK3-targeted proteins, takes a close look at their primary protein sequences, and proposes a model of how MPK3 recognises, binds, and phosphorylates its targets.