Protein and Peptide Letters最新文献

Introduction: Endometrial carcinoma (EC) incidence and mortality continue to rise, and reliable therapeutic targets remain scarce. We aimed to define the oncogenic role and mechanism of tumor protein D52 (TPD52) in EC, focusing on epithelial-mesenchymal transition (EMT) and the PI3K/AKT and ERK/MAPK signaling pathways.

Methods: In this study, we assessed the expression levels of TPD52 in EC tissues and benign endometrial tissues using immunohistochemistry. To further investigate the role of TPD52, we performed experiments both in vitro and in vivo. We transfected siRNA and overexpression (OE) plasmids into Ishikawa and HEC-1-A cell lines to knock down (KD) or overexpress TPD52, respectively. We observed the effects of TPD52 knockdown on tumor growth and EMT through in vitro experiments.

Results: TPD52 was significantly upregulated in EC tissues compared with those of benign endometrial tissues. Silencing TPD52 significantly inhibited cell proliferation, migration, and invasion, whereas TPD52 overexpression produced the opposite effects. TPD52 facilitates epithelial-mesenchymal transition (EMT). Moreover, TPD52 stimulates the PI3K/AKT and ERK/MAPK signaling pathways.

Discussion: These data position TPD52 as a bona fide EC oncoprotein that drives EMT via dual PI3K/AKT-ERK/MAPK signaling. Limitations include the modest patient cohort and the lack of clinical-pathological correlation analyses.

Conclusion: TPD52 promotes EC progression through EMT and PI3K/AKT and ERK/MAPK activation, offering a promising therapeutic target whose clinical utility warrants further investigation.

Introduction: The shepherin II peptide is characterized by a histidine/glycine-rich sequence. This study aimed to design, express recombinantly, and evaluate the antiviral activity of shepherin II against hepatitis A virus (HAV).

Methods: The shepherin II gene was reverse-translated, cloned into the pET-3a vector, and expressed in E. coli BL21 (DE3) pLysS cells induced with 2 mM IPTG. Purification was achieved via cation exchange chromatography, and intact mass analysis using mass spectrometry was carried out. Cytotoxicity on normal Vero cells and antiviral activity on HAV were evaluated.

Results: The mass spectrometry confirmed a primary peptide fragment with a molecular weight of 3,421.30 Da (100% relative abundance). SDS-PAGE verified peptide expression. Cytotoxicity tests on Vero cells showed a CC50 of 219.26 ± 7.91 μg/ml. Antiviral assay revealed an EC50 of 113.92 ± 4.58 μg/ml against HAV, resulting in a selectivity index (SI) of 1.92. This SI indicates limited selectivity compared to the reference drug amantadine, which exhibited an EC50 of 5.67 ± 0.71 μg/ml and an SI of 53.41.

Discussion: The recombinant expression of shepherin II was successfully achieved and confirmed by mass spectrometry and SDS-PAGE. The peptide showed measurable antiviral activity against HAV.

Conclusion: This study demonstrated the feasibility of recombinant shepherin II production and assessed its antiviral activity. However, the limited selectivity index of shepherin II remains a challenge that needs to be addressed through molecular modification or alternative delivery strategies to improve its clinical potential.

The natural horizon of the genetic code has expanded to incorporate amino acids, such as selenocysteine and pyrrolysine. Researchers have incorporated unnatural amino acids (UAAs) into target proteins, demonstrating increased protein functionality depending on their choice and target. The primary challenge in protein engineering is identifying novel antimicrobial short peptides effective against ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.), which are categorized as multidrug-resistant (MDR). UAAs can be preferentially incorporated into short peptides to display therapeutic activity, potentially leading to next-generation targeted therapeutics. In purview of this, we have curated and summarized the applicability of genetic incorporations of UAAs in antimicrobial short peptides with a special emphasis on the importance of green synthesis. The approach affirmed a reduction in the toxicity of peptide drugs, making it biocompatible. This is an efficient protocol to develop novel antimicrobial short peptides catering to precision medications, particularly against MDR pathogens, as a sustainable pharmaceutical approach.

Introduction: The present study aims to compare the monotherapy of diosmetin and 5- hydroxydecanoate (5-HD) against the therapeutic effect of their combination therapy in the unilaterally injected rotenone-induced neurotoxicity in the male rats. Motor deficits accompany Parkinson's Disease (PD), while Bioflavonoids like diosmetin, which are antioxidants and anti-inflammatories, protect against neurotoxins. Moreover, mitochondrial dysfunction contributes to PD. The mitochondrial ATP-sensitive potassium channel [mito(KATP)] regulates reactive species and 5-HD, meaning decreasing it may lessen mitochondrial injury. To evaluate the effect of diosmetin, alone and in combination with 5-HD, on Oxidative Stress (OS) markers, mitochondrial function, and dopaminergic preservation in the SNpc.

Methods: Male Wistar rats were divided into seven groups, including normal control, sham, rotenone-treated, and treatment groups receiving diosmetin, 5-HD, their combination, or selegiline as a standard drug. Biochemical assays were conducted to assess OS markers, mitochondrial complex- I activity, and dopaminergic neuroprotection. Behavioral tests were performed to evaluate motor deficits.

Results: Rotenone administration significantly increased OS, impaired mitochondrial complex-I activity, and reduced motor coordination. Diosmetin treatment significantly reverses the effects of rotenone. Combined treatment with diosmetin and 5-HD showed enhanced neuroprotective effects compared to individual treatments.

Discussion: This study demonstrates that both diosmetin and 5-HD monotherapies alleviate rotenone-induced behavioral impairments in the experimental rats. Additionally, the individual treatment of diosmetin and 5-HD reduces dopaminergic toxicity induced by rotenone. At the sub-- cellular level, diosmetin and 5-HD monotherapies counteract rotenone's impact on antioxidant markers, DA metabolites, and mitochondrial function in the SNpc region of the brain. Notably, combining diosmetin and 5-HD yielded superior therapeutic effects on rotenone-induced behavioral and molecular changes compared to either monotherapy alone. These findings suggest that diosmetin and 5-HD may offer a promising alternative for PD management.

Conclusion: Diosmetin exhibits potent antioxidant and neuroprotective properties against rotenone-induced PD-like pathology. The combination of diosmetin and 5-HD offers a synergistic therapeutic potential, suggesting a promising approach for managing oxidative stress and mitochondrial dysfunction in PD.

Introduction: Numerous X-ray crystal structures of the c-Src SH3 domain have provided a large sampling of atomic-level information for this important signaling domain. Multiple crystal forms have been reported, with variable crystal lattice contacts and chemical crystallization conditions.

Materials and methods: We crystallized the c-Src SH3 domain in a crystallization buffer containing NiCl2.

Results: A unique crystal structure of the Src SH3 domain in the trigonal space group H32 is determined to 1.45 Å resolution. Crystal packing and anomalous scattering reveal that this crystal form is mediated by two ordered nickel ions provided by the crystallization buffer. Nickel coordination occurs in a 2:2 stoichiometry, which dimerizes two SH3 domain monomers across a pseudo-twofold rotation axis and involves the native N-terminal c-Src SH3 amino acid sequence, a surface-exposed histidine residue, and ordered water molecules.

Discussion: This study provides an example of metal-mediated crystallization and metal binding by N-terminal protein residues, contrasting with the Amino-Terminal Copper and Nickel Binding (ATCUN) motif.

Conclusion: Alternative avenues help widen the potential for future crystallography-based studies of the c-Src SH3 domain.

Introduction: Peak-shouldering elution behavior was a common and unexpected result in bind-and-elute mode Cation Exchange Chromatography (CEX), which may be due to the pH transition during the elution step and the aggregation tendency of target proteins.

Methods: Improving the concentration of acid-base pairs in the wash buffers or elution buffers without changing pH or conductivity effectively resolved the peak-shouldering issue in CEX.

Results: In the case of molecule A, the shoulder peak was eliminated in the CEX run by increasing the NaAc-HAc concentration from 50 mM to 100 mM in the elution buffer or from 50 mM to 75 mM in the wash buffer. Higher NaAc-HAc concentrations affect the pH transition in the early stages of the elution step, which may explain the elimination of the shoulder peak. A similar result was observed for molecule B, where increasing the Tris-HCl concentration in the elution buffer from 50 mM to 80 mM also removed the shoulder peak during elution.

Discussion: The successful elimination of peak-shouldering behavior by increasing acid-base pair concentrations highlights the critical role of buffer capacity in modulating pH transitions during CEX. While this strategy offers a simple and effective solution, further investigation is needed to assess its applicability across diverse protein types and buffer systems.

Conclusion: These results demonstrate that increasing the concentration of acid-base pairs in the elution buffer or wash buffer of CEX using NaAc-HAc or Tris-HCl buffers is an effective strategy for eliminating the shoulder-peak.

Introduction: Pleckstrin homology and RhoGEF domain-containing G7 (PLEKHG7) is a largely uncharacterized gene whose role in diffuse large B-cell lymphoma (DLBCL) remains unexplored. Thus, we aimed to profile PLEKHG7 expression, assess its prognostic value, and explore therapeutic implications.

Methods: RNA-seq data from TCGA-DLBCL (n=48) and GTEx normal tissues were analyzed via UCSC XENA. Differential expression was tested using the Wilcoxon rank-sum test and FDR correction. Prognostic significance was evaluated by Kaplan-Meier and multivariate Cox regression (nomogram). Gene set enrichment analysis (GSEA) mapped PLEKHG7-associated pathways. Drug sensitivity correlations were extracted from RNAactDrug. qRT-PCR validated expression in DLBCL cell lines (OCI-Ly3, SU-DHL-4) versus normal B lymphocytes (GM12878).

Results: PLEKHG7 was markedly up-regulated in DLBCL tissues (P < 0.001) and cell lines versus normal controls (AUC = 0.739). High PLEKHG7 expression predicted inferior overall survival (HR = 8.88; 95% CI: 1.09-72.27; P = 0.041) and remained an independent prognostic factor (HR = 10.109; P = 0.033). GSEA linked PLEKHG7 to ribosome, oxidative phosphorylation, proteasome, cytokine-cytokine receptor interaction, spliceosome, and ECM-receptor pathways. Elevated PLEKHG7 negatively correlated with sensitivity to idelalisib, omipalisib, belinostat, methotrexate, and dacinostat.

Discussion: The study's limitations include reliance on bioinformatics data and the lack of functional validation. Further research is needed to elucidate the molecular mechanisms underlying PLEKHG7's role in DLBCL and validate its clinical utility.

Conclusion: PLEKHG7 is significantly overexpressed in DLBCL and independently predicts poor prognosis. Its association with key oncogenic pathways and drug resistance underscores its potential as both a prognostic biomarker and a therapeutic target, warranting further functional validation.

Introduction: Superoxide Dismutases (SODs) are enzymes that catalyze the conversion of toxic free radicals generated during stress conditions into nontoxic forms. Thus, the enzyme superoxide dismutase contributes to the adaptation and survival of microorganisms across a variety of environmental conditions, making it an indispensable enzyme during the response to stress. In this study, we embarked upon investigating and characterizing a Superoxide Dismutase (SOD) from DNA extracted directly from garden soil, where the average temperature ranges from 4°C- 45°C.

Materials and methods: Metagenomic DNA was extracted by employing a kit. The gene was amplified using PCR. The amplified PCR product was gel eluted and ligated into the pGEMT-easy vector and subcloned into an expression vector. The protein was purified using Ni-NTA chromatography and characterized using biophysical, biochemical, and computational approaches.

Results: The recombinant SOD was expressed and purified; the purified protein exhibited activity and stability over a broad pH and temperature range, with optimal activity observed at 40°C and pH 8, respectively. The enzyme remains completely stable at 40°C for 3 h. However, in contrast, it loses 50% of its activity when incubated at 50°C and 60°C for 3 h. The biophysical investigation revealed stable confirmation of the secondary structure of the protein, as evident from circular dichroism and intrinsic Tryptophan (Trp) fluorescence studies. In silico sequence and structural analysis revealed a close similarity of the SOD reported in this study to the Mn SOD of multi- Bacillus species. Molecular simulation dynamics experiments revealed the all-over conformational stability of protein structures at varying pH, indicating broad pH functioning of the enzyme.

Discussion: The study provides a comprehensive analysis of the structure and function of a superoxide dismutase enzyme derived from a soil metagenome. A Mn2+ binding site identified in the study offers an opportunity to further facilitate engineering and design of mutant SOD.

Conclusion: The enzyme exhibits distinct attributes that hold significant industrial relevance. Owing to the wide functionality of SOD at different pH and temperature, it can be tailored for its potential industrial applications, which include its therapeutic potential, thus opening new avenues for enhanced antioxidant therapies and novel biocatalyst designing.

Introduction: The skin of amphibians performs some vital roles, such as camouflage, ion and water transport, and gas exchange. Additionally, it plays a significant role in the immune system by preventing pathogen invasion. The secretions produced by the granular glands in the skin possess antimicrobial properties, which help prevent harmful microorganisms from entering the animal's body. The study aims to determine the total protein amounts in the secretion of Pelophylax bedriagae (Levant water frog) distributed in Türkiye and to reveal whether it has antimicrobial properties. In this context, it is a pioneering study on antimicrobial peptides in the skin secretion of Pelophylax.

Method: Field research was carried out at İzmir, Türkiye's Homeros Valley. Because AMPs are species-specific, not sex-specific peptides, both male and female adults were used, regardless of sex. The average weight of the animals used in the experiment was 21 grams. Granular glands were stimulated using a specialized device, and the secretions were collected for analysis. The collected secretions were lyophilized after centrifugation. Then, the total protein amount in the secretion was determined by Bicinchoninic Acid (BCA). The antibacterial activities of the skin secretions against Escherichia coli and Staphylococcus aureus were assessed using a plate well diffusion assay. The peptide profiles in the skin secretions were determined using the Tricine-SDS-- PAGE electrophoresis method.

Results: The plate well diffusion assay demonstrated that the skin secretions created a 21 mm inhibition zone against E. coli and a 20 mm inhibition zone against S. aureus. The results of the Tricine-SDS-PAGE electrophoresis revealed the presence of peptides with five different molecular weights, including one smaller than 5 kDa. As a result of the analysis, it was determined that P. bedriagae secretion exhibits antimicrobial properties, and many proteins with different molecular masses were identified in the skin secretion.

Discussion: There are no skin secretion studies on P. bedriage in the literature. However, activity studies were also conducted on the skin secretion of another Pelophylax species, P. ridibundus. The study examined the antibacterial activities of the skin secretion against some gram-positive and gram-negative bacteria using a plate well diffusion assay. Their study showed that E. coli formed a 21 mm zone and S. aureus formed a 24 mm zone.

Conclusion: This study is preliminary, and in future studies, AMPs in skin secretions can be isolated by chromatographic methods, such as HPLC, and peptides can be sequenced and identified in detail.

Leukemia is one of the most prevalent malignancies worldwide that causes the unusual evolution of hematopoietic stem cells. The type of leukemia determines the optimal treatment plan and the patient's survival. However, finding safer and more effective medications and developing novel therapeutic strategies are still the most challenging research topics. Nowadays, over half of the medications used to treat cancer are derived from natural ingredients. Medicinal plants are a reliable natural source of anti-leukemic medications. Plant-derived biologically active compounds, including secondary metabolites, have long been considered extremely valuable for treating various human illnesses. However, the limitations of secondary metabolites have led scientists to seek alternative biologically active compounds. Plant-derived proteins and peptides have recently been explored as potential treatments for various human ailments, showing anti-microbial, anti-oxidant, anti-HIV, anti-cancer, ribosome-inactivating, and neuromodulatory properties. Until now, no review article has documented the biologically active proteins and peptides against leukemia. This review article explores the therapeutic properties of plant-derived proteins and peptides against leukemia.