70-kDa Protein Research Articles

Hybrid PET/MRI Imaging of 18F-Fluorodeoxyglucose and 18-kDa Translocator Protein for Presurgical Localization in Refractory Epilepsy.

Surgery remains the only curative option for a third of refractory epilepsy patients, though success depends on precise localization of the epileptogenic zone (EZ). This study aims to assess the clinical value of hybrid 18F-FDG and 18F-DPA-714 PET/MRI for accurate localization and precise boundary delineation. The refractory epilepsy patients who underwent surgery at Xuanwu Hospital from November 2022 to November 2023 were retrospectively recruited. Preoperative simultaneous 18F-FDG and 18F-DPA-714 PET/MRI imaging were analyzed using the asymmetry index (AI) and a 4-point visual score, with the surgical site and pathological findings serving as the gold standard. A total of 43 patients (mean age: 26.30 ± 8.37 years, male: 28) were included in this study. Lesion localization accuracy within the EZ was 76.7% for 18F-FDG PET/MRI, 69.8% for 18F-DPA-714 PET/MRI, and 60.5% for conventional MRI (p = 0.26). In 26 MRI-positive cases, conventional MRI accurately localized all lesions within the EZ, with three cases showing negative findings on 18F-FDG images and six on 18F-DPA-714. Among 17 MRI-negative patients, thirteen demonstrated positive results on hybrid PET/MRI. Additionally, 18F-DPA-714 PET/MRI proved more effective in delineating lesion boundaries. Compared to 18F-FDG, the AI score was significantly lower (0.25 ± 0.18 vs. 0.46 ± 0.19, p < 0.001), while the visual score was higher (4.00 ± 2.00 vs. 3.00 ± 0.00, p = 0.01). 18F-DPA-714 PET/MRI can effectively complement conventional MRI in the preoperative assessment of refractory epilepsy, with localization accuracy on par with 18F-FDG and enhanced capability in delineating lesion boundaries.

Determination of Urinary Neutrophil Gelatinase-Associated Lipocalin (uNGAL) Reference Intervals in Healthy Adult and Pediatric Individuals Using a Particle-Enhanced Turbidimetric Immunoassay.

Background: The current gold standards for diagnosing acute kidney injury (AKI) are an increase in serum creatinine and a decrease in urine output, which are inadequate for rapid diagnosis. Neutrophil gelatinase-associated lipocalin (NGAL) is a 25-kDa protein produced and secreted by injured kidney tubule epithelial cells, and can serve as an early urinary biomarker for AKI. ProNephro AKI (NGAL) is an immunoassay for the quantitative determination of NGAL in urine (uNGAL) that recently received FDA clearance. A multisite, cross-sectional study was conducted to establish reference intervals for uNGAL in apparently healthy individuals. Methods: Urine samples were collected from apparently healthy individuals aged ≥3 months who met all inclusion criteria and no exclusion criteria. Specimens were temporarily stored at room temperature or 2-8 °C, then transferred into urinalysis tubes before being frozen and shipped for testing. uNGAL testing was performed using the ProNephro AKI (NGAL) immunoassay on a Roche cobas c501 analyzer. Results: Of the 688 individuals screened, 677 were eligible, and 629 (91.4%) of those were deemed evaluable. The 95th and 97.5th percentile uNGAL values for all pediatric participants were below the clinical cutoff of 125 ng/mL. uNGAL values were statistically significantly higher for female vs. male participants in both adult (p = 0.003) and pediatric groups (p < 0.001), while differences were not statistically significant for age, site location, race, or ethnicity. Conclusions: This study provides normal reference intervals for uNGAL with the ProNephro AKI (NGAL) clinical chemistry immunoassay that may be useful for interpreting patient results.

First-in-Human Study of [11C]NCGG401 for Imaging Colony-Stimulating Factor 1 Receptors in the Brain.

Microglia, the immune cells in the brain, play a significant role in the pathophysiology of neurodegenerative diseases. To visualize these cells in the living brain, we developed a PET ligand, [11C]NCGG401 (4-{2-[((1R,2R)-2-hydroxycyclohexyl)(methyl)amino]benzothiazol-6-yloxy}-N-methylpicolinamide, NCGG401), that targets colony-stimulating factor 1 receptor (CSF1R). In this study, we present the first-in-human evaluation of [11C]NCGG401 to assess its safety profile and then to evaluate its kinetics to quantify CSF1R in the human brain. Methods: Head to upper thigh PET scans were conducted in 3 healthy men to estimate the effective dose of [11C]NCGG401. Brain PET scans were performed on 6 healthy men, combined with arterial blood sampling and metabolite analyses. Compartmental and graphical models were used to quantify CSF1R in the human brain. [11C]NCGG401 PET data were indirectly compared with regional CSF1R protein levels after death that were reported in a proteomics study. In addition, the results of this study were directly compared with the PET imaging of 18-kDa translocator protein using [11C]DPA-713 (N,N-diethyl-2-[2-(4-methoxyphenyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidin-3-yl]acetamide, DPA-713). Results: The administration of [11C]NCGG401 did not result in severe adverse events. The effective doses per injected activity were 5.1 ± 0.2 µSv/MBq for men and 6.1 ± 0.3 µSv/MBq for women. [11C]NCGG401 demonstrated good brain permeability, with peak uptake reaching an SUV of 3. Regional total distribution volumes were reliably quantified using the 2-tissue compartment model and a Logan plot with 60 min of scan data. The resulting parametric images reflected the known distribution of CSF1R in the brain. Furthermore, regional total distribution volume values of [11C]NCGG401 showed good correlation with regional CSF1R protein levels. The [11C]NCGG401 images showed regional distributions different from those of [11C]DPA-713. Conclusion: [11C]NCGG401 images appear to reflect regional microglia-specific distributions of CSF1R in the brain, consistent with the findings of a CSF1R proteomics study by others. However, ultimate confirmation of specific CSF1R binding should be validated by evaluating, in suitable preclinical or human experiments, pharmacologic blockade of its binding in the brain invivo.

LysR-Type Transcriptional Regulator VirR Responds to Temperature and pH and Directly Activates the Transcription of virS-Containing Operon in Rhodococcus equi.

Rhodococcus equi-a facultative intracellular pathogen of macrophages-causes bronchopneumonia in foals and patients who are immunocompromised. Virulent strains of R. equi possess a virulence-associated plasmid, which encodes a 15- to 17-kDa surface protein called virulence-associated protein A (VapA). VapA expression is regulated by temperature and pH. Two transcriptional regulators, VirR and VirS, are involved in the transcriptional regulation of vapA. VirR regulates VapA expression through VirS. However, whether VirR directly regulates virS transcription is unclear. In this study, we examined VirR binding to the promoter region of the icgA operon, which contains virS, using the electrophoretic mobility shift assay and DNase I footprinting. VirR bound DNA fragments containing the virR-icgA intergenic region. Transcription from the promoter in this region was VirR-dependent and regulated by temperature and pH. The VirR-binding site contained the LysR-type transcriptional regulator-binding consensus motif, T-N11-A. A point mutation (L98E) in the putative ligand-binding pocket of VirR constitutively activated the icgA promoter. However, no apparent difference was observed in the electrophoretic mobility shift assay and DNase I footprinting using the icgA promoter when L98E VirR was compared with wild-type VirR. A bacterial two-hybrid system identified an interaction between VirR and RpoA. Our data reveal that VirR binds the promoter of the icgA operon and directly activates its transcription. Furthermore, the regulation of VapA expression in response to temperature and pH is mediated by VirR.

The new anti-angiogenesis perspective of rheumatoid arthritis with geniposide: Reducing the extracellular release of HSP70 in HUVECs.

Angiogenesis is essential for pannus formation and maintenance in rheumatoid arthritis (RA). Heat shock protein 70kDa (HSP70) can induce angiogenesis by being released extracellularly through exosomes. Geniposide (GE) is the primary pharmacological component of the fruit of Gardenia jasminoides Ellis (GJ). In vivo, we have found that GE is able to reduce HSP70 levels in the synovium and serum of CIA-S and has anti-angiogenic effects. However, the mechanism by which GE inhibits HSP70 to improve angiogenesis is still unclear. This study aims to explore how GE inhibits the extracellular release of HSP70 and its impact on angiogenesis in human umbilical vein endothelial cells (HUVECs). HUVECs' exosomes were extracted using ultracentrifugation and characterized through transmission electron microscope, nanoparticle tracer technology, nano-flow cytometry and Western blotting. The proliferative ability of HUVECs was assessed by EdU and CCK8 assay. Transwell and wound healing assays were used to measure the migration ability of HUVECs, while tube formation assay was employed to evaluate their tube-forming ability. The TNF-α-induced HSP70 release model in HUVECs was established, with extracellular HSP70 levels serving as an evaluation index. Immunofluorescence and co-immunoprecipitation assay were used to analyze the interaction between HSP70 and the lipid raft marker Caveolin-1 (Cav-1). Western blotting was employed to investigate the expression of SphK1/S1P/S1PRs/Gαi pathway-related proteins, and ELISA was utilized to detect extracellular S1P and HSP70 levels. The exosomes of HUVECs contained HSP70. HUVECs were stimulated by extracellular HSP70, which enhanced their proliferation, migration, and tube-forming abilities. TNF-α (10ng/mL) significantly increased the release of HSP70, which was inhibited by GE (25µM-100µM) in a concentration-dependent manner. GE reduced HSP70 in lipid rafts without affecting Cav-1. GE (100µM) inhibited proteins in the SphK1/S1P/S1PRs/Gαi pathway, preventing HSP70 release and improving HUVECs' functions compared to the K6PC-5 (SphK1-specific agonist) and TNF-α groups. This study found that GE inhibited the extracellular release of HSP70 by suppressing the SphK1/S1P/S1PRs/Gαi pathway, thereby producing anti-angiogenic effects in vitro. This provides a novel direction and strategy for anti-angiogenesis therapy for RA.

BCL-2 Protein Expression in Epithelial Ovarian Cancer: Association with Age and Histopathological Features in Uganda

Background: Globally, ovarian cancer accounted for 314,000 new cases and 207,000 deaths in 2020. In Uganda, 626 cases were documented over 25 years, with an average incidence rate of 7.1 per 100,000 women. Epithelial ovarian carcinoma is the predominant subtype, and BCL-2, a 26-kDa anti-apoptotic protein, has been associated with chemotherapy resistance. However, no study has assessed BCL-2 expression in epithelial ovarian cancer in Uganda. Objective: This study aimed to determine the prevalence of BCL-2 protein expression in epithelial ovarian cancer and its association with age and histopathological features in Uganda. Methods: A retrospective cross-sectional study was conducted using randomly sampled archived tissue blocks from 113 patients diagnosed with epithelial ovarian cancer. Samples underwent hematoxylin and eosin (H&amp;E) and immunohistochemistry (IHC) staining to evaluate BCL-2 expression. Results: The mean patient age was 55 years, with the largest proportion (36.3%) aged 50–59 years. BCL-2 expression was observed in 36.3% of cases. There was no statistically significant association between BCL-2 expression and age (p = 0.396), histological subtype (p = 0.520), or tumor grade (p = 0.833). High-grade serous carcinoma was the predominant histological subtype (46%), and well-differentiated carcinoma was the most common grade (53%). Conclusion: BCL-2 expression was prevalent in epithelial ovarian cancer in Uganda but showed no significant association with age, histological grade, or subtype. These findings underscored the need for further research to explore the prognostic and therapeutic implications of BCL-2 in ovarian cancer management.

Isolation and characterization of bacteriophages targeting methicillin-resistant Staphylococcus aureus (MRSA) from burn patients and sewage water: a genomic and proteomic study.

The spectrum of infections caused by methicillin-resistant Staphylococcus aureus (MRSA) ranges from minor conditions to potentially life-threatening diseases. The rising antibiotic resistance in MRSA often leads to treatment failures, underscoring the urgent need for novel eradication strategies. This study focuses on isolating MRSA from burn patients, determining its antibiogram profile, and isolating and characterizing bacteriophages from sewage water that target MRSA, alongside conducting genomic analysis of the phages. A total of 70 samples were collected from burn patients, with MRSA identification and characterization performed using a combination of biochemical and molecular techniques, as well as antibiotic sensitivity testing. Based on host range analysis, a specific phage (phage-3) was selected for detailed characterization, including proteomic analysis, genetic mapping, phylogenetic studies, and analysis of open reading frames (ORFs) and motifs. The prevalence of MRSA in the samples was found to be 28.6%. Antibiotic susceptibility tests indicated that 94% of the MRSA isolates were sensitive to tobramycin and gentamicin, while vancomycin exhibited the lowest sensitivity, with only 2% effectiveness. Using the soft agar overlay method, three bacteriophages (phage-1, phage-2, and phage-3) were successfully isolated from sewage water. Among these, phage-3 exhibited the broadest host range. Further analysis showed that phage-3 demonstrated optimal activity at pH levels between 6 and 8, and within a temperature range of 20-40°C. Phage-3 also displayed a rapid adsorption phase within the first 0-5min, and its one-step growth curve revealed a latent period lasting up to 30min, followed by a significant increase in titer from 30 to 50min. Proteomic analysis of phage-3 identified the presence of 33kDa and 65kDa proteins. Phylogenetic analysis showed that phage-3 shares 96.6% similarity with Mammallicoccus phage vB_MscM-PMS3. The ORF analysis identified 80 potential ORFs within the phage's entire genome.

Current advances in the structure-activity relationship (SAR) analysis of the old/new 18-kDa translocator protein ligands.

The translocator protein 18kDa (TSPO) is a crucial external mitochondrial protein involved in cholesterol translocation, which is essential for steroid production. As a primary marker of neuroinflammation, TSPO has been implicated in the development and progression of various neurodegenerative and neuropsychiatric disorders. This review highlights the structural diversity of TSPO ligands, many of which have undergone modifications from selective central benzodiazepine receptor (CBR) ligands to enhance their affinity for TSPO. The paper discusses the significant advancements in the design of these ligands, emphasizing their binding efficacy and specificity. Additionally, it provides an update on the progress of several TSPO ligands that have advanced to clinical trials. The review aims to elucidate the structure-activity relationships (SAR) that govern the interaction between TSPO and its ligands, thereby offering insights into the development of new therapeutic agents targeting TSPO for the treatment of neuroinflammatory conditions. Overall, this work provided an update on previous finding and serves as a valuable resource for researchers in the field.

Conformational change induced by binding of Mn2+ ions activates SloR transcription factor in Streptococcus mutans.

Streptococcus mutans, a bacterium commonly found in the human oral cavity, is considered the primary causative agent of dental caries. A key player in the pathophysiology of S. mutans is SloR, a 25-kDa metalloregulatory protein. SloR plays a crucial role in coordinating the uptake of essential metal ions, particularly manganese, with the transcription of the bacterium's virulence genes. To elucidate the molecular mechanism underlying the enhanced binding affinity of SloR to DNA upon Mn2+ ion binding, a combination of computational (QM and MD) and experimental (ITC, DSC, CD, EPR) methods have been employed. Computational simulations revealed that Mn2+ binding induces a conformational change of SloR, primarily affecting the positioning of its DNA-binding domains, bringing them to an appropriate position for DNA binding. Consequently, the protein's DNA binding affinity is modulated. Additionally, experimental findings indicate that the SloR monomer binds up to three Mn2+ ions and that the thermodynamic stability of SloR increases upon Mn2+ complexation. The presented computational results also suggest that Mn2+ binding at the primary binding sites is sufficient to trigger the observed conformational change in SloR.

Anti-aquaporin-5 and anti-poly-U-binding-factor-60kDa protein antibodies in primary Sjögren's disease patients: preliminary data and correlation with disease activity indices.

In primary Sjögren's disease (pSjD), in addition to glandular inflammation and atrophy, functional secretion impairment may contribute to dryness. Altered protein distribution and antibodies against aquaporin-5 (anti-AQP5) and poly-U-binding factor 60kDa protein (anti-PUF60) have been reported in pSjD and may be specifically implicated in the glandular secretive processes. This study aimed to assess the occurrence of serum anti-AQP5 and anti-PUF60 antibodies and their correlations with clinical and laboratory features of pSjD. Blood samples from pSjD patients and healthy donors (HD) were collected, and anti-AQP5 and anti-PUF60 antibodies were detected using an enzyme-linked immunosorbent assay. Differences between groups were evaluated using appropriate statistical tests, and odds ratios (OR) of high disease activity were assessed by multivariate stepwise backward multiple regression and adjusted for clinical covariates. Serum samples from 36 pSjD patients and 8 HD were analysed, and anti-AQP5 and anti-PUF60 antibody levels were not significantly different between groups. However, pSjD patients with high disease activity (n. 10) had significantly higher levels of anti-AQP5 antibodies compared to those with low-moderate disease activity (p<0.001). At logistic regression analysis, variables associated with high disease activity were anti-AQP5 (OR 128.9, 95% CI 2.7-615), C-reactive protein (OR 12.9, 95% CI 1.2-137.2), and C4 <10 mg/dl (OR 60, 95% CI 1.1-318.9). Our pilot study confirms that anti-AQP5 antibodies may discriminate pSjD patients with high disease activity. These findings offer valuable clinical implications for managing pSjD patients, potentially identifying patients at high risk of glandular deterioration.

Systemic Granulomatosis in the Meagre Argyrosomus regius: Fishing for a Plausible Etiology.

Meagre (Argyrosomus regius) is one of the fast-growing species considered for sustainable aquaculture development along the Mediterranean and Eastern Atlantic coasts. The emergence of Systemic Granulomatosis (SG), a disease marked by multiple granulomas in various tissues, poses a significant challenge in meagre aquaculture. In the current study, we investigate the association of Mycobacterium spp. and SG in offshore aquaculture facilities in Sardinia, Italy. A total of 34 adult seemingly healthy meagre were arbitrarily collected and analyzed, combining histological, microbiological, molecular, metagenomics, and in situ techniques to investigate the presence of pathogens. Ziehl-Neelsen (ZN), periodic acid-schiff (PAS), and Giemsa stains were performed for the detection of acid-fast bacteria, common parasites, and fungi within granulomas, respectively. Granulomas were detected in 91% (31/34) of fish. The affected organs were kidney (88%), liver (47%), heart (41%), intestine (17.6%), and brain (5%). Acid-fast staining, along with Mycobacterium spp. specific quantitative PCR (qPCR), in situ hybridization (ISH) assay, and microbiological analyses showed negative results for the detection of Mycobacterium spp. and other bacteria implicated in granuloma formation. However, PCR amplification and sequencing of the 65-kDa heat shock protein gene revealed the presence of M. chelonae in 13% of both formalin-fixed and frozen liver tissues. Bacterial isolation failed to detect nontuberculous mycobacteria (NTM) and other bacteria typically associated with granulomas. Consistently, the use of an M. chelonae-specific probe in ISH failed to identify this bacterial species in granulomas. Collectively, results do not support the role of M. chelonae in the development of granulomas and suggest rejecting the hypothesis of a potential link between NTM and SG.

Insights into the regulation of CHIP E3 ligase-mediated ubiquitination of neuronal protein BNIP-H.

BCL2/adenovirus E1B 19-kDa protein-interacting protein 2 homolog (BNIP-H or Caytaxin), a pivotal adaptor protein that facilitates cerebellar cortex growth and synaptic transmission, is posttranslationally modified to regulate neuronal function. This study reports the ubiquitination of BNIP-H by Carboxyl terminus of Hsc70-Interacting Protein (CHIP), a U-box containing E3 ligase that is also regulated via autoubiquitination. Specifically, it was observed that CHIP autoubiquitinated itself primarily at Lys23 and Lys31 in vitro. Mutation of these residues shows the autoubiquitination of successive lysines of CHIP. In total, nine lysines on CHIP were identified as the autoubiquitination sites, the collective mutation of which almost completely terminated its autoubiquitination. Additionally, CHIP-mediated ubiquitination of BNIP-H is completely inhibited when BNIP-H bears arginine mutations at four key lysine residues. Next, using hydrogen deuterium exchange mass spectrometry, a model of a plausible mechanism was proposed. The model suggests transient N-terminal interactions between the CHIP and BNIP-H which allows for the swinging of U-box domain of CHIP to ubiquitinate BNIP-H. Following complex dissociation, BNIP-H population is regulated via the ubiquitin-proteasome pathway. Collectively, these results aid in our understanding of CHIP-mediated BNIP-H ubiquitination and provide further insight into the roles of these proteins in neuritogenesis and neurotransmission.

Reference intervals for serum prolactin concentrations: analytical and clinical considerations.

Prolactin is a 23kDa protein synthesised and released by lactotrophs located in the anterior pituitary gland. The main function of prolactin is the development of the mammary glands and the production and maintenance of milk secretion during pregnancy and lactation. Prolactin is present in 3 forms in the circulation monomeric, dimeric and macroprolactin which is one of the causes of variability between assay manufacturers. Although most assays have been standardised using the WHO's 3rd international standard there is still considerable variability between commercially available assays and as such laboratories need to establish their own reference intervals. Clinicians use reference intervals to distinguish between healthy and diseased individuals. Direct methods to establish reference intervals are expensive and time consuming and are impractical for most routine clinical laboratories. Indirect methods using the laboratories own data are more commonly used but also have drawbacks. Determination of reference intervals for prolactin is particularly difficult due to various physiological causes for variation in prolactin levels including age, sex and menopause. The presence of macroprolactin, its detection and methods for reporting its presence adds to reference interval determination complexity and harmonisation of these methods may improve clinical care.

Genetic diversity and phylogenetic relationships of Theileria equi inferred from heat shock protein 70 kDa gene sequences in Brazilian horses.

Equine piroplasmosis, a tick-borne disease caused by hemoparasites of the Babesia and Theileria genera, has significant clinical and economic impacts worldwide. This study aims to characterize the heat shock protein 70kDa (hsp70) gene sequences of Theileria equi from naturally infected horses across the five geographic regions in Brazil, and to analyze the phylogenetic relationships between T. equi and other parasites within the order Piroplasmida. Primers specific to T. equi were designed through in silico analysis of hsp70 gene sequences available in GenBank. Forty samples of equine whole blood were subjected to DNA extraction and conventional PCR targeting the hsp70 gene. The resulting amplicons were purified and sequenced. The Hsp70 sequences generated were aligned using the ClustalW method, and phylogenetic analyses were conducted using Bayesian Inference (BI) and Maximum Likelihood (ML) methods. The evolutionary distance within and between species of parasites from the order Piroplasmida was evaluated, as was the entropy of the hsp70 gene sequence among Theileria species. Phylogenetic analysis of the forty sequences identified in this study revealed two circulating genotypes of T. equi in Brazil. One genotype comprised the American sequence along with thirty-six Brazilian sequences, while the other grouped a sequence from Mongolia with four Brazilian sequences. The Equus clade was distinctly placed with strong support, indicating a different evolutionary ancestor compared to Theileria spp. and Babesia spp. groups. The results confirmed hsp70 as an effective molecular marker for phylogeny, yielding topologies similar to those observed with 18S rDNA while distinguishing closely related protozoan species such as T. haneyi and T. equi. This study provides valuable evolutionary insights into the phylogenetic relationships within the order Piroplasmida.

Salivary heat shock protein 70 as a potential non-invasive biomarker of environmental thermal stress in dairy cattle.

The present study aims to explore the potential biomarker application of salivary heat shock 70kDa protein in detecting thermal stress in dairy animals noninvasively. The study spans for 45 days during the mid-summer season (April-May), involving twelve multiparous non-pregnant adult Jersey crossbred cows by randomly allocating them into groups (six animals in each group). The control animals were maintained in the shed, whereas the thermal stress group animals were exposed to environment heat between 10:00h to 16.00h and they were feed and watered ad libitum. During the experimental period, the hematobiochemical, physiological, behavioural, nutritional and production responses were recorded and the whole blood and saliva were collected fortnightly. Results revealed significant increase in WBC, AST, ALP, blood urea nitrogen, triglycerides, cholesterol, HDL, blood and salivary cortisol, respiratory rate, rectal temperature, skin temperature of neck, lumbar and forelimb regions, standing time, salivary and blood HSP70 mRNA expression and their protein concentrations in heat stressed animals. In addition, RBC, haemoglobin, MCV, PCV, platelet, platelet-large cell ratio (PLCR), lying time, feed intake, milk yield and rumination time were significantly decreased in thermally stress animals. Furthermore, ROC curve analysis revealed the biomarker potential of these significantly altered parameters with 100% sensitivity and specificity for predicting environmental heat stress in dairy cows with AUC and Youden's - index of 1.00 except platelet. Moreover, salivary HSP70 demonstrated significant correlation with these biomarkers. Noteworthily, salivary HSP70 also exhibited strong association with blood HSP70 and salivary cortisol. Furthermore, salivary HSP70 revealed 100% sensitivity and specificity in discriminating the dairy cattle succumbed to heat stress from healthy. In conclusion, the present study provides a newer insight into the multifaceted roles of HSP70 and identified salivary heat shock 70kDa protein as a potential, reliable and more sensitive non-invasive biomarker for identifying environmental heat stress in dairy cattle.

Homology Modelling and Molecular Docking of Some Natural Compounds as Inhibitors of Anopheles gambiae Heat Shock Protein 70KDa and Bifunctional Glutamyl/prolyl-tRNA Synthetase

Homology Modelling and Molecular Docking of Some Natural Compounds as Inhibitors of Anopheles gambiae Heat Shock Protein 70KDa and Bifunctional Glutamyl/prolyl-tRNA Synthetase

Prolactin: structure, receptors, and functions.

Prolactin (PRL) is a 23-kDa protein synthesized and secreted by lactotroph cells of the anterior pituitary gland but also by other peripheral tissues. PRL binds directly to a unique transmembrane receptor (PRLR), and the JAK2/signal transducer and activator of transcription 5 (Stat5) pathway is considered the major downstream pathway for PRLR signaling. To a lesser extent, PRL may be cleaved into the shorter 16-kDa PRL, also called vasoinhibin, whose signaling is not fully known. According to rodent models of PRL signaling inactivation and the identification of human genetic alterations in PRL signaling, a growing number of biological processes are partly mediated by PRL or its downstream effectors. In this review, we focused on PRL structure and signaling and its canonical function in reproduction. In addition to regulating reproductive functions, PRL also plays a role in behavior, notably in initiating nurturing and maternal behavior. We also included recent insights into PRL function in several fields, including migraines, metabolic homeostasis, inflammatory and autoimmune disease, and cancer. Despite the complexity of understanding the many functions of PRL, new research in this field offers interesting perspectives on physiological and pathophysiological processes.

Recombinant expression, downstream optimization, and therapeutic evaluation of recombinant human interleukin-37 for cancer therapy.

Interleukin-37 is a cytokine with potent immunosuppressive properties that has been shown to have potential to treat autoimmune and chronic inflammatory diseases, as well as certain types of cancer. IL-37 is a 19kDa protein which interacts with proteins in receptor-dependent and receptor-independent pathways. The expression of the IL-37 protein cloned into the pET-28a vector was optimized in Rosetta 2(DE3) after comparing its expression with Rosetta-gami 2(DE3) and Rosetta 2(DE3) pLysS, which was then used for the large-scale production of IL-37. IMAC purification of IL-37 yielded > 97% pure 0.9mg/mL protein from auto-induced fermentation. The IC50 value of IL-37 was < 1µM, which was similar to that of doxorubicin, and proliferation of > 80% of all cancer cells was inhibited by 100µg/mL of IL-37 protein. IL-37 may be a promising theragnostic target for cancer due to its comparable IC50 value with that of doxorubicin.

One-step nanoscale expansion microscopy reveals individual protein shapes.

The attainable resolution of fluorescence microscopy has reached the subnanometer range, but this technique still fails to image the morphology of single proteins or small molecular complexes. Here, we expand the specimens at least tenfold, label them with conventional fluorophores and image them with conventional light microscopes, acquiring videos in which we analyze fluorescence fluctuations. One-step nanoscale expansion (ONE) microscopy enables the visualization of the shapes of individual membrane and soluble proteins, achieving around 1-nm resolution. We show that conformational changes are readily observable, such as those undergone by the ~17-kDa protein calmodulin upon Ca2+ binding. ONE is also applied to clinical samples, analyzing the morphology of protein aggregates in cerebrospinal fluid from persons with Parkinson disease, potentially aiding disease diagnosis. This technology bridges the gap between high-resolution structural biology techniques and light microscopy, providing new avenues for discoveries in biology and medicine.

Allelic Diversity and Development of Breeder-Friendly Marker Specific to floury2 Gene Regulating the Accumulation of α-Zeins and Essential Amino Acids in Maize Kernel.

Maize zeins lack essential amino acids, such as methionine, lysine, and tryptophan. The floury2 (fl2) mutation reduces zein synthesis and increases methionine and lysine content in kernels. In this study, fl2 gene (1612bp) was sequenced in eight wild-type and two mutant inbreds and detected 218 SNPs and 18 InDels. Transversion of C to T at 343bp position caused the substitution of alanine by valine in the fl2 mutant. A PCR-based marker (FL-SNP-CT) was developed, which distinguished the favorable mutant fl2 allele (T) from the wild-type (C) Fl2 allele. Gene-based diversity analysis using seven gene-based InDel markers grouped 48 inbred lines into three major clusters, with an average genetic dissimilarity coefficient of 0.534. The average major allele frequency, gene diversity, heterozygosity, and polymorphism information content of the InDel markers were 0.701, 0.392, 0.039, and 0.318, respectively. Haplotype analysis revealed 29 haplotypes of fl2 gene among these 48 inbreds. Amino acid substitution (Ala-Val) at the signal peptide cleavage site produced unprocessed 24-kDa mutant protein instead of 22-kDa zein found in normal genotype. Eight paralogues of fl2 detected in the study showed variation in exon lengths (616-1170bp) and translation lengths (135-267 amino acids). Orthologue analysis among 15 accessions of Sorghum bicolor and two accessions of Saccharum spontaneum revealed a single exon in fl2 gene, ranging from 267 to 810bp. The study elucidated the molecular basis of fl2 mutation and reported a breeder-friendly marker for molecular breeding programs. This is the first study to characterize fl2 gene in a set of subtropically adapted inbreds.