Small Peptides Research Articles

The BCL11A transcription factor stimulates the enzymatic activities of the OGG1 DNA glycosylase.

The BCL11A transcription factor has previously been shown to interact with and stimulate the enzymatic activities of the NTHL1 DNA glycosylase and Pol β polymerase. Here we show that BCL11A and a smaller peptide encompassing amino acids 160 to 520 can interact with the 8-oxoguanine DNA glycosylase, OGG1, increase the binding of OGG1 to DNA that contains an 8-oxoguanine base and stimulate the glycosylase activity of OGG1. Following BCL11A knockdown, we observed an increase in oxidized purines in the genome using comet assays, while immunoassays reveal an increase in 8-oxoG bases. Structure-function analysis indicates that the stimulation of OGG1 by BCL11A requires the zinc fingers 1, 2 and 3 as well as the proline-rich region between the first and second zing finger, but a glutamate-rich region downstream of zinc finger 3 is dispensable. Ectopic expression of a small peptide that contains the three zinc fingers can rescue the increase in 8-oxoguanine caused by BCL11A knockdown. These findings, together with previous results showing that BCL11A stimulates the enzymatic activities of NTHL1 and the Pol β polymerase, suggest that high expression of BCL11A is important to protect cancer cells against oxidative DNA damage.

PCP-bε is a novel positive regulator of pollen germination in Arabidopsis thaliana

small cysteine-rich peptides play essential roles in different stages of the plant reproductive process. Pollen germination is a prerequisite for double fertilization and is directly related to seed formation and crop yield. However, the small cysteine-rich peptides that are involved in pollen germination remain to be identified. In this study, identification and phylogenetic analysis of PCP-Bε genes in sequenced Brassicaceae show that pollen coat protein B-class protein PCP-Bε gene is widespread in Arabidopsis and its high relatives, but lost in some Brassica species. Expression analyses display that AtPCP-Bε gene is expressed in Arabidopsis pollen. Arabidopsis PCP-Bε knockout mutants are generated by CRISPR/Cas9, Phenotypic analyses show that the absence of AtPCP-Bε obviously impairs in vitro pollen germination, but has no influence on pollen tube growth, which demonstrates that AtPCP-Bε is a novel positive regulator of pollen germination. It is speculated that AtPCP-Bε should interact with the receptor from pollen to perform its function. These findings are useful for further analysis on the molecular mechanism of pollen germination.

Multi-Omic Approaches in Cancer-Related Micropeptide Identification.

Despite the advances in modern cancer therapy, malignant diseases are still a leading cause of morbidity and mortality worldwide. Conventional treatment methods frequently lead to side effects and drug resistance in patients, highlighting the need for novel therapeutic approaches. Recent findings have identified the existence of non-canonical micropeptides, an additional layer of the proteome complexity, also called the microproteome. These small peptides are a promising class of therapeutic agents with the potential to address the limitations of current cancer treatments. The microproteome is encoded by regions of the genome historically annotated as non-coding, and its existence has been revealed thanks to recent advances in proteomic and bioinformatic technology, which dramatically improved the understanding of proteome complexity. Micropeptides have been shown to be biologically active in several cancer types, indicating their therapeutic role. Furthermore, they are characterized by low toxicity and high target specificity, demonstrating their potential for the development of better tolerated drugs. In this review, we survey the current landscape of known micropeptides with a role in cancer progression or treatment, discuss their potential as anticancer agents, and describe the methodological challenges facing the proteome field of research.

Techniques, Databases and Software Used for Studying Polar Metabolites and Lipids of Gastrointestinal Parasites.

Gastrointestinal parasites (GIPs) are organisms known to have coevolved for millennia with their mammalian hosts. These parasites produce small molecules, peptides, and proteins to evade or fight their hosts' immune systems and also to protect their host for their own survival/coexistence. The small molecules include polar compounds, amino acids, lipids, and carbohydrates. Metabolomics and lipidomics are emerging fields of research that have recently been applied to study helminth infections, host-parasite interactions and biochemicals of GIPs. This review comprehensively discusses metabolomics and lipidomics studies of the small molecules of GIPs, providing insights into the available tools and techniques, databases, and analytical software. Most metabolomics and lipidomics investigations employed LC-MS, MS or MS/MS, NMR, or a combination thereof. Recent advancements in artificial intelligence (AI)-assisted software tools and databases have propelled parasitomics forward, offering new avenues to explore host-parasite interactions, immunomodulation, and the intricacies of parasitism. As our understanding of AI technologies and their utilisation continue to expand, it promises to unveil novel perspectives and enrich the knowledge of these complex host-parasite relationships.

Research on Bitter Peptides in the Field of Bioinformatics: A Comprehensive Review.

Bitter peptides are small molecular peptides produced by the hydrolysis of proteins under acidic, alkaline, or enzymatic conditions. These peptides can enhance food flavor and offer various health benefits, with attributes such as antihypertensive, antidiabetic, antioxidant, antibacterial, and immune-regulating properties. They show significant potential in the development of functional foods and the prevention and treatment of diseases. This review introduces the diverse sources of bitter peptides and discusses the mechanisms of bitterness generation and their physiological functions in the taste system. Additionally, it emphasizes the application of bioinformatics in bitter peptide research, including the establishment and improvement of bitter peptide databases, the use of quantitative structure-activity relationship (QSAR) models to predict bitterness thresholds, and the latest advancements in classification prediction models built using machine learning and deep learning algorithms for bitter peptide identification. Future research directions include enhancing databases, diversifying models, and applying generative models to advance bitter peptide research towards deepening and discovering more practical applications.

Intermedin Alleviates Diabetic Cardiomyopathy by Up-Regulating CPT-1β through Activation of the Phosphatidyl Inositol 3 Kinase/Protein Kinase B Signaling Pathway.

Diabetic cardiomyopathy (DCM), one of the most serious long-term consequences of diabetes, is closely associated with myocardial fatty acid metabolism. Carnitine palmitoyltransferase-1β (CPT-1β) is the rate-limiting enzyme responsible for β-oxidation of long-chain fatty acids. Intermedin (IMD) is a pivotal bioactive small molecule peptide, participating in the protection of various cardiovascular diseases. However, the role and underlying mechanisms of IMD in DCM are still unclear. In this study, we investigated whether IMD alleviates DCM via regulating CPT-1β. A rat DCM model was established by having rats to drink fructose water for 12 weeks. A mouse DCM model was induced by feeding mice a high-fat diet for 16 weeks. We showed that IMD and its receptor complexes levels were significantly down-regulated in the cardiac tissues of DCM rats and mice. Reduced expression of IMD was also observed in neonatal rat cardiomyocytes treated with palmitic acid (PA, 300 μM) in vitro. Exogenous and endogenous IMD mitigated cardiac hypertrophy, fibrosis, dysfunction, and lipid accumulation in DCM rats and IMD-transgenic DCM mice, whereas knockout of IMD worsened these pathological processes in IMD-knockout DCM mice. In vitro, IMD alleviated PA-induced cardiomyocyte hypertrophy and cardiac fibroblast activation. We found that CPT-1β enzyme activity, mRNA and protein levels, and acetyl-CoA content were increased in T2DM patients, rats and mice. IMD up-regulated the CPT-1β levels and acetyl-CoA content in T2DM rats and mice. Knockdown of CPT-1β blocked the effects of IMD on increasing acetyl-CoA content and on inhibiting cardiomyocyte hypertrophy and cardiac fibroblast activation. IMD receptor antagonist IMD17-47 and the phosphatidyl inositol 3 kinase (PI3K)/protein kinase B (Akt) inhibitor LY294002 reversed the effects of IMD on up-regulating CPT-1β and acetyl-CoA expression and on inhibiting cardiomyocyte hypertrophy and cardiac fibroblast activation. We revealed that IMD alleviates DCM by up-regulating CPT-1β via calcitonin receptor-like receptor/receptor activity-modifying protein (CRLR/RAMP) receptor complexes and PI3K/Akt signaling. IMD may serve as a potent therapeutic target for the treatment of DCM.

Comprehensive characterization of the rhizosphere metabolome: A novel method for non-targeted analysis of Lotus japonicus root exudates

Plant root exudates play a pivotal role in shaping soil dynamics and the microbial community in the rhizosphere. The chemical composition of root exudates includes primary and secondary metabolites, including amino acids, organic acids, flavonoids, and small peptides. Comprehensive characterization of root exudates will allow for a better understanding of rhizosphere processes and interactions, but analysis of root exudates is hindered by complicated collection setups, time-consuming sample preparation, and a lack of definitive annotations within metabolomics. We present a method optimized for non-targeted analysis of primary and secondary metabolites in root exudate samples using ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry. The method was tested on root exudates from Lotus japonicus, collected using distinct and well-established sampling methods: a hydroponic-soil-hybrid approach, as well as a modification of a soil-leaching method, thus exemplifying the versatility of the analysis method. The method allows for non-targeted screening of plant metabolites, and provides low detection limits (0.002–0.05 μg/mL) and high recoveries (78 ± 30%), though a matrix effect was observed for certain plant metabolites. Detection of a large number of features was achieved (670–2785) of which the majority could be putatively annotated at the compound class level. Of these, 14 features were putatively annotated to a specific structure with high confidence, three of which were confirmed with analytical reference standards. The method can be used for investigation of the overall change in root exudation, as well as for investigating significant changes in metabolites in response to intraenous and extraneous parameters.

When can we trust structural models derived from pair distribution function measurements?

The pair distribution function (PDF) is an important metric for characterising structure in complex materials, but it is well known that meaningfully different structural models can sometimes give rise to equivalent PDFs. In this paper, we discuss the use of model likelihoods as a general approach for discriminating between such homometric structure solutions. Drawing on two main case studies-one concerning the structure of a small peptide and the other amorphous calcium carbonate-we show how consideration of model likelihood can help drive robust structure solution, even in cases where the PDF is particularly information-poor. The obvious thread of these individual case studies is the potential role for machine-learning approaches to help guide structure determination from the PDF, and our paper finishes with some forward-looking discussion along these lines.

Fabrication of Poly Lactic-co-Glycolic Acid Microneedles for Sustained Delivery of Lipophilic Peptide-Carfilzomib.

Transdermal drug delivery (TDD) is an attractive route of administration, providing several advantages, especially over oral and parenteral routes. However, TDD is significantly restricted due to the barrier imposed by the uppermost layer of the skin, the stratum corneum (SC). Microneedles is a physical enhancement technique that efficiently pierces the SC and facilitates the delivery of both lipophilic and hydrophilic molecules. Dissolving microneedles is a commonly used type that is fabricated utilizing various biodegradable and biocompatible polymers, such as polylactic acid, polyglycolic acid, or poly(lactide-co-glycolide) (PLGA). Such polymers also promote the prolonged release of the drug due to the slow degradation of the polymer matrix following its insertion. We selected carfilzomib, a small therapeutic peptide (MW: 719.924 g/mol, log P 4.19), as a model drug to fabricate a microneedle-based sustained delivery system. This study is a proof-of-concept investigation in which we fabricated PLGA microneedles using four types of PLGA (50-2A, 50-5A, 75-5A, and 50-7P) to evaluate the feasibility of long-acting transdermal delivery of carfilzomib. Micromolding technique was used to fabricate the PLGA microneedles and characterization tests, including Fourier transform infrared spectroscopy, insertion capability using the skin simulant Parafilm model, histological evaluation, scanning electron microscopy, and confocal microscopy were conducted. In vitro release and permeation testing were conducted in vertical Franz diffusion cells. N-methyl pyrrolidone was utilized as the organic solvent and microneedles were solidified in controlled conditions, which led to good mechanical strength. Both in vitro release and permeation testing showed sustained profiles of carfilzomib over 7 days. The release and permeation were significantly influenced by the molecular weight of PLGA and the lipophilic properties of carfilzomib.

Structure, Function, and Activity of Small Molecule and Peptide Inhibitors of Protein Arginine Methyltransferase 1.

Protein arginine N-methyltransferases (PRMT) are a family of S-adenosyl-l-methionine (SAM)-dependent enzymes that transfer methyl-groups to the ω-N of arginyl residues in proteins. PRMTs are involved in regulating gene expression, RNA splicing, and other activities. PRMT1 is responsible for most cellular arginine methylation, and its dysregulation is involved in many cancers. Accordingly, many groups have targeted PRMT1 using small molecules and peptide inhibitors. In this Perspective, we discuss the structure and function of selected peptide and small molecule inhibitors of PRMT1. We examine inhibitors that target the substrate arginyl peptide, SAM, or both binding sites, and the type of inhibition that results. Small molecules, and peptides that are bisubstrate, and/or PRMT transition state mimic inhibitors as well as inhibitors that alkylate PRMTs will be discussed. We define a structure-activity relationship for the aromatic/heteroaromatic N-methylethylenediamine inhibitors of PRMT1 and review current progress of PRMT1 inhibitors in clinical trials.

Promising Therapeutic Strategies for Hematologic Malignancies: Innovations and Potential.

In this review we explore innovative approaches in the treatment of hematologic cancers by combining various therapeutic modalities. We discuss the synergistic potential of combining inhibitors targeting different cellular pathways with immunotherapies, molecular therapies, and hormonal therapies. Examples include combining PI3K inhibitors with proteasome inhibitors, NF-κB inhibitors with immunotherapy checkpoint inhibitors, and neddylation inhibitors with therapies targeting the tumor microenvironment. Additionally, we discuss the potential use of small molecules and peptide inhibitors in hematologic cancer treatment. These multidimensional therapeutic combinations present promising strategies for enhancing treatment efficacy and overcoming resistance mechanisms. However, further clinical research is required to validate their effectiveness and safety profiles in hematologic cancer patients.

Ways to reduce radioactivity accumulation in the kidney during targeted therapy using small molecules, peptides and antibody fragments

Administration of pharmaceuticals containing radioactive isotopes and capable of specific binding to certain proteins is one of the approaches used in the treatment or diagnosis of malignant tumors. High renal accumulation of radioactive compounds after administration of radioconjugates with molecular mass less than 70 KDa is of the challenges that need to be solved. The purpose of the study was to identify the most effective approaches to reduce the accumulation of radioactivity in the kidneys after administration of radioconjugates used for diagnostic imaging and targeted therapy for cancer. Material and Methods. We conducted a literature search on the topic of the review in the electronic databases PubMed, Scopus and Web of Science from 1987 to 2023, 82 articles were used for writing the review. Results. The review presents a description of approaches used to improve the biodistribution of radioconjugates, mainly in preclinical studies. The advantages and disadvantages of such techniques have been described. Conclusion. Reducing renal radioactivity using radioconjugates of molecules with molecular masses less than 70 KDa is a challenging but achievable task. It is concluded that the use of cleavable linkers in such radioconjugates is highly promising, since this approach does not change the pharmacokinetics of such drugs. It is noted that the advantage of introducing concomitant substances compared to changing the structure of radioconjugates is a lesser dependence on the characteristics of a particular radiopharmaceutical. This approach also does not require prior work to modify the radioconjugate, but has limited efficiency.

Recent approaches in the application of antimicrobial peptides in food preservation.

Antimicrobial peptides (AMPs) are small peptides existing in nature as an important part of the innate immune system in various organisms. Notably, the AMPs exhibit inhibitory effects against a wide spectrum of pathogens, showcasing potential applications in different fields such as food, agriculture, medicine. This review explores the application of AMPs in the food industry, emphasizing their crucial role in enhancing the safety and shelf life of food and how they offer a viable substitute for chemical preservatives with their biocompatible and natural attributes. It provides an overview of the recent advancements, ranging from conventional approaches of using natural AMPs derived from bacteria or other sources to the biocomputational design and usage of synthetic AMPs for food preservation. Recent innovations such as structural modifications of AMPs to improve safety and suitability as food preservatives have been discussed. Furthermore, the active packaging and creative fabrication strategies such as nano-formulation, biopolymeric peptides and casting films, for optimizing the efficacy and stability of these peptides in food systems are summarized. The overall focus is on the spectrum of applications, with special attention to potential challenges in the usage of AMPs in the food industry and strategies for their mitigation.

Experimental Investigation on Electrical Conductivity Variation of Carnosine and Zinc Chloride Aqueous Solutions under Microwave Irradiation.

The mechanism of biological effects of environmental electromagnetic radiation is still not completely clear. The chelation of biological small molecule peptides with metal ions plays a very important role in human metabolism. In this paper, a special experimental system was designed to measure the conductivity of carnosine and zinc chloride mixed aqueous solutions under different concentration ratios, microwave powers, and temperatures. The experimental results show that, first, different concentration ratios can alter the conductivity change rate of the mixed aqueous solution. The conductivity of the solution always increases under microwave irradiation at a concentration ratio of 1:1. However, the conductivity is reduced by -0.04% with a 1:5 concentration ratio and 6 W microwave power at 10 °C. Second, temperature can alter the conductivity change rate of the aqueous mixture. The higher the temperature, the smaller the conductivity change rate. Third, different microwave powers can alter the conductivity change rate of the mixed aqueous solution. In general, the conductivity change rate increases with an increase in microwave power. Experimentally observed reduction of the conductivity change rate in carnosine and zinc chloride aqueous solution under low microwave power and low temperature indicates that microwaves do affect the chelation of carnosine with zinc chloride. This work provides a new perspective for the mechanism of explanation of microwave biological effects.

Formation of Nanofibrillar Self-Healing Hydrogels Using Antimicrobial Peptides.

The rise of drug-resistant microorganisms has prompted the development of innovative strategies with the aim of addressing this challenge. Among the alternative approaches gaining increased attention are antimicrobial peptides (AMPs), a group of peptides with the ability to combat microbial pathogens. Here, we investigated a small peptide, KLVFF, derived from the Alzheimer's amyloid-β (Aβ) protein. While Aβ has been associated with the development of neurodegenerative diseases, the core part of the Aβ protein, namely the Aβ 16-20 fragment, has also been exploited to obtain highly functional biomaterials. In this study we found that KLVFF is capable of self-assembling into a fibrillar network to form a self-healing hydrogel. Moreover, this small peptide can undergo a transition from a gel to a liquid state following application of shear stress, in a reversible manner. As an AMP, this material exhibited both antibacterial and antifungal properties while remaining highly biocompatible and noncytotoxic toward mammalian cells. The propensity of the KLVFF hydrogel to rapidly assemble into highly ordered macroscopic structures makes it an ideal candidate for biomedical applications necessitating antimicrobial activity, such as wound healing.

First-Generation Radiolabeled Cyclic Peptides for Molecular Imaging of Platelet-Derived Growth Factor Receptor α.

Occult nodal spread and metastatic disease require longstanding imaging and biochemical assessments for thyroid cancer, a disease that has a propensity for diffuse, small-volume disease. We have developed a 64Cu-labeled platelet-derived growth factor receptor α (PDGFRA) antibody for immuno-PET of PDGFRA in metastatic papillary thyroid cancer (PTC). The present work describes the discovery of small cyclic PDGFRA-targeting peptides, their binding features, and radiolabeling with positron emitter gallium-68 (68Ga) for in vitro and in vivo characterization in thyroid cancer models. Phage-display technology with two separate libraries and seven different cell lines was used through three rounds of biopanning as well as flow cytometry and comparative analysis with recombinant protein to select specific peptide sequences. Phenotypic binding analysis was completed by using phosphorylation and cell migration assays. In vitro protein binding was analyzed with thermophoresis and flow cytometry using the fluorescent-labeled PDGFRA peptide. Peptide candidates were modified with the NOTA chelator for radiolabeling with 68Ga. In vitro cell uptake was studied in various thyroid cancer cell lines. In vivo studies of 68Ga-labeled peptides included metabolic stability and PET imaging. From the original library (1013 compounds), five different peptide groups were identified based on biopanning experiments with and without the α subunit of PDGFR, leading to ∼50 peptides. Subsequent phenotypic screening revealed two core peptide sequences (CP16 and CP18) that demonstrated significant changes in the level of PDGFRA phosphorylation and cell migration. Alanine scan sublibraries were created from these two lead peptide sequences, and peptides were radiolabeled using 68Ga-GaCl3 at pH 4.5, resulting in RCP > 95% within 34-40 min, including SPE purification. Cyclic peptide CP18.5 showed the strongest effects on cell migration, flow cytometry, and binding by visual interference color assay. 68Ga-labeled PDGFRA-targeting peptides showed elevated cell and tumor uptake in models of thyroid cancer, with 68Ga-NOTA-CP18.5 being the lead candidate. However, metabolic stability in vivo was compromised for 68Ga-NOTA-CP18.5 vs 68Ga-NOTA-CP18 but without impacting tumor uptake or clearance profiles. First-generation radiolabeled cyclic peptides have been developed as novel radiotracers, particularly 68Ga-NOTA-CP18.5, for the molecular imaging of PDGFRA in thyroid cancer.

The anti-inflammatory and tolerogenic potential of small spleen peptides.

Maintaining peripheral immune tolerance and preventing harmful autoimmune reactions is a fundamental task of the immune system. However, these essential functions are significantly compromised during autoimmune disorders, creating a major challenge in treating these conditions. In this context, we provide an overview of research on small spleen polypeptides (SSPs) that naturally regulate peripheral immune tolerance. Alongside outlining the observed effects of SSPs, we summarize here the findings on the cellular and molecular mechanisms that underlie their regulatory impact. Specifically, SSPs have demonstrated remarkable effectiveness in halting the progression of developing or established autoimmune disorders like psoriasis or arthritis in animal models. They primarily target dendritic cells (DCs), swiftly prompting the production of extracellular ATP, which is then degraded and sensed by adenosine receptors. This process triggers the mTOR signaling cascade, similar to powerful immune triggers, but instead of a rapid and intense reaction, it leads to a moderate yet significant activation of the mTOR signaling cascade. This induces a tolerogenic state in dendritic cells, ultimately leading to the generation of Foxp3+ immunosuppressor Treg cells. In addition, SSPs may indirectly attenuate the autoimmune response by reducing extracellular ATP synthesis in non-immune cells, such as endothelial cells, when exposed to elevated levels of proinflammatory cytokines. SSPs thus have the potential to contribute to the restoration of peripheral immune tolerance and may offer valuable therapeutic benefits in treating autoimmune diseases.

Co-culturing Bifidobacterium animalis ssp. lactis with Lactobacillus helveticus accelerates its growth and fermentation in milk through metabolic interactions

[Objective] This study aimed to investigate the interaction between Lactobacillus helveticus H9 (H9) and Bifidobacterium animalis ssp. lactis Probio-M8 (M8) through metabolomics analysis, focusing on understanding how co-culturing these strains can enhance bacterial growth and metabolism, thereby shortening the fermentation cycle and improving efficiency. [Methods] The H9 and M8 strains were cultured individually and in combination (1:1 ratio) in milk. The fermented milk metabolomes were analyzed using solid-phase microextraction-gas chromatography-mass spectrometry. [Results] In the dual-strain fermentation, the M8 strain exhibited a 2.33-fold increase in viable bacterial count compared with single-strain fermentation. Additionally, the dual-strain fermentation resulted in greater metabolite abundance and diversity. Notably, the dual-strain fermented milk showed significantly elevated levels of metabolites, including 5-methyl-2-hexanone, (E)-3-octen-2-one, acetic acid, alanine, and 3-hydroxy-butanal. [Conclusion] Our results demonstrated that co-culturing the M8 and H9 strains accelerated growth and fermentation efficiency. This enhancement effect is likely attributed to the strong proteolytic ability of the H9 strain, which hydrolyzes casein to produce small molecular peptides, alanine, tyrosine, and other growth-promoting factors. The insights gained from this study have significant implications for probiotics and the dairy industry, potentially leading to shorter fermentation cycles, enhanced cost-effectiveness, and improved nutritional and functional properties of future fermented milk products. Additionally, these findings may contribute to advancements in probiotic research and applications.

Targeting proliferating cell nuclear antigen enhances ionizing radiation-induced cytotoxicity in prostate cancer cells.

Proliferating cell nuclear antigen (PCNA) is essential for DNA replication and repair, cell growth, and survival. PCNA also enhances androgen receptor (AR) signaling in prostate cancer (PC) cells. We identified a PCNA interaction protein (PIP) box at the N-terminal domain of AR and developed a small peptide PCNA inhibitor R9-AR-PIP containing AR PIP-box. We also identified a series of small molecule PCNA inhibitors (PCNA-Is) that bind directly to PCNA and interrupt PCNA functions. The present study investigated the effects of the PCNA inhibitors on the sensitivity of PC cells to X-ray radiation. The effects of targeting PCNA on radio sensitivity of PC cells were investigated in four lines of castration-resistant PC (CRPC) cells with different AR expression statuses. The cells were treated with the PCNA inhibitors and X-ray radiation alone or in combination. The effects of the treatment on expression of AR target genes, DNA damage response, DNA damage, homologous recombination repair (HRR), and cytotoxicity were evaluated. We found that the androgen response element (ARE) occupancy of the DNA damage response gene PARP1 by AR is significantly attenuated by PCNA-I1S or R9-AR-PIP combined with X-ray radiation, while X-ray radiation alone does not enhance the ARE occupancy. PCNA-I1S or R9-AR-PIP alone significantly inhibits occupancy of the AR-occupied regions (AROR) in PRKDC and XRCC2 genes. R9-AR-PIP and PCNA-I1S inhibit expression of AR-Vs target gene cyclin A2 and show the additive effects with radiation in AR-positive CRPC cells. Targeting PCNA by PCNA-I1S and R9-AR-PIP downregulates expression of DNA damage response genes EXO1, Rad54L, Rad51, and/or PARP1 and shows the additive effects with radiation as compared with their respective controls in AR-positive CRPC LNCaP-AI, 22Rv1, and R1-D567 cells, but not in AR-negative PC-3 cells. R9-AR-PIP and PCNA-I1S elevate the levels of phospho-DNA-PKcs(S2056) and γH2AX, indicating DNA damage in response to radiation in AR-positive cells. The HRR is significantly attenuated by PCNA inhibitors PCNA-I1S, R9-AR-PIP, and T2AA in all four CRPC cells examined, and inhibited by Enzalutamide (Enz) only in 22RV1 cells. The cytotoxicity induced by X-ray radiation in androgen-dependent LNCaP cells is enhanced by Enz and a lower concentration of R9-AR-PIP in the colony formation assay. R9-AR-PIP at higher concentration reduces the colony formation and has an additive effect with X-ray radiation in all AR expressing cells, regardless of AR-FL and AR-Vs, but does not significantly alter the colony formation in AR-negative PC-3 cells. PCNA-I1S attenuates colony formation and has an additive effect with ionizing radiation in all four CRPC cells, regardless of AR expression status. These data provide a strong rationale for the therapy studies using PCNA-I1S or R9-AR-PIP in combination with X-ray radiation against CRPC tumors in preclinical models.

Screening and characterization of umami peptides from enzymatic and fermented products of wheat gluten using machine learning

To explore the umami peptides from wheat gluten hydrolysates (WGHs) with different degrees of hydrolysis (DH) after Corynebacterium glutamicum (C. glutamicum) fermentation, a high-throughput screening method for umami peptides was established based on machine learning. The results showed fermented WGHs with high DHs exhibited more favourable taste profile after 4 days of fermentation and produced more small molecule peptides. Fermentation increased the number of characteristic peptides in WGHs with DH10, DH15, and DH20. Among, fermented WGHs with DH15 were the important source of potential umami peptides. Six potential umami peptides were screened (QQLPQFEE, LSFE, EELR, YTCE, YTTD, EEDQ) and were demonstrated to form stable complex with the T1R1/T1R3 receptor through hydrogen bonding interactions, electrostatic interactions, and hydrophobic interactions forces. The affinity of peptides and receptor were in the range of −6.3 to −8.1 kcal/mol. GLU735, GLY755, SER734, SER757 and THR227 were the critical binding sites. Sensory evaluation and electronic tongue showed that six peptides exhibited umami profile and had the capacity to enhance umami and saltiness taste. EEDQ presented the strongest intensity of umami taste and QQLPQFEE had the lowest umami thresholds. This study provides reliable methods for effectively screening new microbial-derived umami peptides.