Identification Of Proteins Research Articles

Navigating the Neurobiology of Parkinson’s: The Impact and Potential of α-Synuclein

Parkinson’s disease (PD) is the second most prevalent neurodegenerative disease worldwide; therefore, since its initial description, significant progress has been made, yet a mystery remains regarding its pathogenesis and elusive root cause. The widespread distribution of pathological α-synuclein (αSyn) aggregates throughout the body raises inquiries regarding the etiology, which has prompted several hypotheses, with the most prominent one being αSyn-associated proteinopathy. The identification of αSyn protein within Lewy bodies, coupled with genetic evidence linking αSyn locus duplication, triplication, as well as point mutations to familial Parkinson’s disease, has underscored the significance of αSyn in initiating and propagating Lewy body pathology throughout the brain. In monogenic and sporadic PD, the presence of early inflammation and synaptic dysfunction leads to αSyn aggregation and neuronal death through mitochondrial, lysosomal, and endosomal functional impairment. However, much remains to be understood about αSyn pathogenesis, which is heavily grounded in biomarkers and treatment strategies. In this review, we provide emerging new evidence on the current knowledge about αSyn’s pathophysiological impact on PD, and its presumable role as a specific disease biomarker or main target of disease-modifying therapies, highlighting that this understanding today offers the best potential of disease-modifying therapy in the near future.

Abstract A067: Identifying and targeting FOLFIRINOX resistance mechanisms to improve outcomes in pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with a 5-year survival rate of only 12%. More than 80% of PDAC patients cannot undergo potentially curative surgical resection due to the presentation of advanced, metastatic disease, and are instead offered systemic chemotherapy regimens to which resistance is rapidly acquired. FOLFIRINOX chemotherapy offers an 11.1-month median survival compared to 6.8 months with Gemcitabine. Rapid onset of resistance to FOLFIRINOX, coupled with the lack of alternative targeted therapies, ultimately results in patients succumbing to the disease. My project aims to identify mechanisms by which PDAC cells acquire resistance to FOLFIRINOX treatment and investigate interventions aimed at enhancing and/or reinstating FOLFIRINOX sensitivity. Our lab has propagated distinct patient derived xenografts (PDXs) from the Avner Australian Pancreatic Cancer Matrix Atlas (APMA) and I have exposed them to 11 - 15 rounds of FOLFIRINOX (or vehicle control) in vivo to reflect the clinical management of PDAC. Additionally, intrapancreatic orthotopic tumors derived from cells from the KPC mouse model (LSL-KrasG12D/+;LSL-Trp53R172H/+;Pdx-1-Cre) have received 5 - 6 rounds of FOLFIRINOX (or vehicle control). These tumors will undergo quantitative data-independent acquisition mass spectrometry proteomic analysis and RNA sequencing transcriptomic analysis (RNAseq) to reveal resistance pathways and/or proteins operating in the chronically treated tumors and the surrounding pancreatic tumor microenvironment. Additionally, we have developed and cultured matched treatment naïve and FOLFIRINOX-treated patient-derived cell lines (PDCLs) and mouse cell lines from the PDX and KPC-derived tumors. These lines will allow us to validate future targets via gene knockdown and/or pharmacological intervention. 3D assays, such as our organotypic matrix models, will be performed to assess the metastatic potential of cancer cells and their response to FOLFIRINOX in vitro. Furthermore, subcutaneous and orthotopic (intrapancreatic) models coupled with intravital (in vivo) imaging and biosensor technology will allow us to assess targeting in live tumors and metastatic sites in real time. It is the hope that this research will lead to the identification of proteins or protein pathways which can be manipulated in order to re-sensitize tumors to FOLFIRINOX chemotherapy. Citation Format: Victoria Tyma, Ying-Fei Liew, Alice Tran, Anna Howell, Shona Ritchie, Avner Australian Pancreatic Cancer Matrix Atlas, Australian Pancreatic Genome Initiative, Thomas Cox, Marina Pajic, David Herrmann, Brooke Pereira, Paul Timpson, Katie Gordon. Identifying and targeting FOLFIRINOX resistance mechanisms to improve outcomes in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A067.

In-Depth Host Cell Protein Analysis and Viral Protein Impurity Monitoring in Adeno-Associated Virus-Based Gene Therapy Products Using Optimized Wide Window Data-Dependent Acquisition Method.

Compared to other protein therapeutics, there is currently limited knowledge about the residual host cell proteins (HCPs) in adeno-associated virus (AAV)-based gene therapy products. This is primarily due to the lack of a robust and sensitive mass spectrometry-based method for HCP analysis in AAV samples. Existing liquid chromatography-mass spectrometry methods used for analyzing HCPs in therapeutic monoclonal antibodies (mAbs) often cannot be directly applied to AAVs, due to some unique characteristics of AAV samples encountered during their development such as limited sample availability/protein concentration and the presence of surfactants. In this study, we have developed a novel workflow for robust and in-depth HCP analysis of AAV samples by combining wide-window data-dependent acquisition for improved low-abundance HCP detection with single-pot, solid-phase-enhanced sample preparation (SP3) for low-input sample preparation. Using this newly developed method, we were able to detect more than 650 HCPs in a commercial AAV1 sample with a high quantitative reproducibility. This represents a greater than 5-fold increase in HCP protein identification compared to an in-solution digestion method followed by traditional data-dependent acquisition. Similar benefits can also be achieved for other AAV serotypes that were produced internally and purified through different processes. The detection limit of this method is as low as 0.06 ng/mL, enabling more comprehensive HCP coverage in AAV samples. Moreover, for the first time, we have identified several process-related viral proteins, such as Rep 78 and E4. These proteins need to be closely monitored during AAV process development as they may present a greater risk for immunogenicity compared to HCPs that are derived from human HEK293 cells.

Genome-wide identification of CaWD40 proteins reveals the involvement of a novel complex (CaAN1-CaDYT1-CaWD40-91) in anthocyanin biosynthesis and genic male sterility in Capsicum annuum

BackgroundThe WD40 domain, one of the most abundant in eukaryotic genomes, is widely involved in plant growth and development, secondary metabolic biosynthesis, and mediating responses to biotic and abiotic stresses. WD40 repeat (WD40) protein has been systematically studied in several model plants but has not been reported in the Capsicum annuum (pepper) genome.ResultsHerein, 269, 237, and 257 CaWD40 genes were identified in the Zunla, CM334, and Zhangshugang genomes, respectively. CaWD40 sequences from the Zunla genome were selected for subsequent analysis, including chromosomal localization, phylogenetic relationships, sequence characteristics, motif compositions, and expression profiling. CaWD40 proteins were unevenly distributed on 12 chromosomes, encompassing 19 tandem duplicate gene pairs. The 269 CaWD40s were divided into six main branches (A to F) with 17 different types of domain distribution. The CaWD40 gene family exhibited diverse expression patterns, and several genes were specifically expressed in flowers and seeds. Yeast two-hybrid (Y2H) and dual-luciferase assay indicated that CaWD40-91 could interact with CaAN1 and CaDYT1, suggesting its involvement in anthocyanin biosynthesis and male sterility in pepper.ConclusionsIn summary, we systematically characterized the phylogeny, classification, structure, and expression of the CaWD40 gene family in pepper. Our findings provide a valuable foundation for further functional investigations on WD40 genes in pepper.

Profiling sorghum-microbe interactions with a specialized photoaffinity probe identifies key sorgoleone binders in Acinetobacter pittii.

Interactions between plants and soil microbial communities that benefit plant growth and enhance nutrient acquisition are driven by the selective release of metabolites from plant roots, or root exudation. To investigate these plant-microbe interactions, we developed a photoaffinity probe based on sorgoleone (sorgoleone diazirine alkyne for photoaffinity labeling, SoDA-PAL), a hydrophobic secondary metabolite and allelochemical produced in Sorghum bicolor root exudates. We applied SoDA-PAL to the identification of sorgoleone-binding proteins in Acinetobacter pittii SO1, a potential plant growth-promoting microbe isolated from sorghum rhizosphere soil. Competitive photoaffinity labeling of A. pittii whole cell lysates with SoDA-PAL identified 137 statistically enriched proteins, including putative transporters, transcriptional regulators, and a subset of proteins with predicted enzymatic functions. We performed computational protein modeling and docking with sorgoleone to prioritize candidates for experimental validation and then confirmed binding of sorgoleone to four of these proteins in vitro: the α/β fold hydrolase SrgB (OH685_09420), a fumarylacetoacetase (OH685_02300), a lysophospholipase (OH685_14215), and an unannotated hypothetical protein (OH685_18625). Our application of this specialized sorgoleone-based probe coupled with structural bioinformatics streamlines the identification of microbial proteins involved in metabolite recognition, metabolism, and toxicity, widening our understanding of the range of cellular pathways that can be affected by a plant secondary metabolite.IMPORTANCEHere, we demonstrate that a photoaffinity-based chemical probe modeled after sorgoleone, an important secondary metabolite released by sorghum roots, can be used to identify microbial proteins that directly interact with sorgoleone. We applied this probe to the sorghum-associated bacterium Acinetobacter pittii and showed that probe labeling is dose-dependent and sensitive to competition with purified sorgoleone. Coupling the probe with proteomics and computational analysis facilitated the identification of putative sorgoleone binders, including a protein implicated in a conserved pathway essential for sorgoleone catabolism. We anticipate that discoveries seeded by this workflow will expand our understanding of the molecular mechanisms by which specific metabolites in root exudates shape the sorghum rhizosphere microbiome.

MiLoPYP: self-supervised molecular pattern mining and particle localization in situ

AbstractCryo-electron tomography allows the routine visualization of cellular landscapes in three dimensions at nanometer-range resolutions. When combined with single-particle tomography, it is possible to obtain near-atomic resolution structures of frequently occurring macromolecules within their native environment. Two outstanding challenges associated with cryo-electron tomography/single-particle tomography are the automatic identification and localization of proteins, tasks that are hindered by the molecular crowding inside cells, imaging distortions characteristic of cryo-electron tomography tomograms and the sheer size of tomographic datasets. Current methods suffer from low accuracy, demand extensive and time-consuming manual labeling or are limited to the detection of specific types of proteins. Here, we present MiLoPYP, a two-step dataset-specific contrastive learning-based framework that enables fast molecular pattern mining followed by accurate protein localization. MiLoPYP’s ability to effectively detect and localize a wide range of targets including globular and tubular complexes as well as large membrane proteins, will contribute to streamline and broaden the applicability of high-resolution workflows for in situ structure determination.

Identification of STAM-binding protein as a target for the treatment of gemcitabine resistance pancreatic cancer in a nutrient-poor microenvironment

Pancreatic cancer (PC) is a highly malignant solid tumor whose resistance to gemcitabine (GEM) chemotherapy is a major cause of poor patient prognosis. Although PC is known to thrive on malnutrition, the mechanism underlying its chemotherapy resistance remains unclear. The current study analyzed clinical tissue sample databases using bioinformatics tools and observed significantly upregulated expression of the deubiquitinase STAMBP in PC tissues. Functional experiments revealed that STAMBP knockdown remarkably increases GEM sensitivity in PC cells. Multiple omics analyses suggested that STAMBP enhances aerobic glycolysis and suppresses mitochondrial respiration to increase GEM resistance in PC both in vitro and in vivo. STAMBP knockdown decreased PDK1 levels, an essential regulator of the aerobic glycolytic process, in several cancers. Mechanistically, STAMBP promoted the PDK1-mediated Warburg effect and chemotherapy resistance by modulating E2F1 via direct binding to E2F1 and suppressing its degradation and ubiquitination. High-throughput compound library screening using three-dimensional protein structure analysis and drug screening identified the FDA drug entrectinib as a potent GEM sensitizer and STAMBP inhibitor, augmenting the antitumor effect of GEM in a patient-derived xenograft (PDX) model. Overall, we established a novel mechanism, via the STAMBP–E2F1–PDK1 axis, by which PC cells become chemoresistant in a nutrient-poor tumor microenvironment.

Component-resolved diagnosis: enhancing precision diagnosis and clinical management of shellfish allergies

Shellfish, being one of the eight major food allergens, affects approximately 3% of the global population. The occurrence of shellfish allergy is not only related to the individual's immune system sensitivity but is also influenced by geographical environment, food availability, and dietary habits. Although crustaceans (such as shrimp, crab, and lobster) and mollusks (such as oysters, mussels, and squid) are collectively referred to as shellfish, they exhibit significant differences in biological evolution and the spectrum of allergenic molecules they contain, leading to various allergic reactions. Accurate identification of allergenic proteins is crucial for the diagnosis and management of shellfish allergies, with key allergenic protein families including tropomyosin, arginine kinase, and hemocyanin. Furthermore, due to the diversity of shellfish allergens and their cross-reactivity with dust mite and insect allergens, diagnosing and managing shellfish allergies is complex, especially concerning tropomyosin and arginine kinase protein families. Currently, there are no specific immunotherapy treatments for shellfish allergies, and clinical management primarily relies on avoiding allergens and using anti-allergy medications. This article thoroughly interprets the " Molecular Allergology User's Guide 2.0 (MAUG 2.0)" published by the European Academy of Allergy and Clinical Immunology (EAACI) and the latest research on shellfish allergies both domestically and internationally. It highlights the significant role of allergen component diagnostics in optimizing the diagnostic and treatment processes for shellfish allergies, effectively assisting clinicians in accurately identifying common allergens and cross-reactions, thereby providing patients with more personalized diagnosis and treatment plans.

Slot Blot- and Electrospray Ionization-Mass Spectrometry/Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry-Based Novel Analysis Methods for the Identification and Quantification of Advanced Glycation End-Products in the Urine.

Proteins, saccharides, and low molecular organic compounds in the blood, urine, and saliva could potentially serve as biomarkers for diseases related to diet, lifestyle, and the use of illegal drugs. Lifestyle-related diseases (LSRDs) such as diabetes mellitus (DM), non-alcoholic steatohepatitis, cardiovascular disease, hypertension, kidney disease, and osteoporosis could develop into life-threatening conditions. Therefore, there is an urgent need to develop biomarkers for their early diagnosis. Advanced glycation end-products (AGEs) are associated with LSRDs and may induce/promote LSRDs. The presence of AGEs in body fluids could represent a biomarker of LSRDs. Urine samples could potentially be used for detecting AGEs, as urine collection is convenient and non-invasive. However, the detection and identification of AGE-modified proteins in the urine could be challenging, as their concentrations in the urine might be extremely low. To address this issue, we propose a new analytical approach. This strategy employs a method previously introduced by us, which combines slot blotting, our unique lysis buffer named Takata's lysis buffer, and a polyvinylidene difluoride membrane, in conjunction with electrospray ionization-mass spectrometry (ESI)/matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS). This novel strategy could be used to detect AGE-modified proteins, AGE-modified peptides, and free-type AGEs in urine samples.

Genome-Wide Mapping of RNA-Protein Associations via Sequencing.

RNA-protein interactions are crucial for regulating gene expression and cellular functions, with their dysregulation potentially impacting disease progression. Systematically mapping these interactions is resource-intensive due to the vast number of potential RNA and protein interactions. Here, we introduce PRIM-seq ( P rotein- R NA Interaction M apping by sequencing), a method for the concurrent de novo identification of RNA-binding proteins (RBPs) and the elucidation of their associated RNAs. PRIM-seq works by converting each RNA-protein pair into a unique chimeric DNA sequence, which is then decoded through DNA sequencing. Applied to two human cell types, PRIM-seq generated a comprehensive human RNA-protein association network (HuRPA), consisting of more than 350,000 RNA-proteins pairs involving approximately 7,000 RNAs and 11,000 proteins. The data revealed an enrichment of previously reported RBPs and RNA-protein interactions within HuRPA. We also identified LINC00339 as a protein-associating non-coding RNA and PHGDH as an RNA-associating protein. Notably, PHGDH interacts with BECN1 and ATF4 mRNAs, suppressing their protein expression and consequently inhibiting autophagy, apoptosis, and neurite outgrowth while promoting cell proliferation. PRIM-seq offers a powerful tool for discovering RBPs and RNA-protein associations, contributing to more comprehensive functional genome annotations.

Identification of proteins’ expression pathway and the effective miRNAs for the treatment of human papillomavirus-induced cervical cancer: in-silico analyses-experimental research

Introduction: Cervical cancer is the fourth most common cancer in women. The risk factors for cervical cancer include human papillomavirus (HPV) infection, age, smoking, number of pregnancies, use of oral contraceptives, and diet. However, long-term HPV infection appears to be the main risk factor for developing cervical cancer. This in silico analysis aims to identify the expression network of proteins and the miRNAs that play a role in the development of HPV-induced cervical cancer. Methods: The critical proteins and miRNAs were extracted using the DisGeNET and miRBase databases. String and Gephi were applied to the network analysis. The GTEx web tool was utilized to Identify tissue expression levels. The Enrichr website was used to explore the molecular function and pathways of found genes. Results: Ten proteins, TP53, MYC, AKT1, TNF, IL6, EGFR, STAT3, CTNNB1, ESR1, and JUN, were identified as the most critical shared gene network among cervical cancer and HPV. Seven miRNAs were found, including hsa-mir-146a, hsa-mir-27, hsa-mir-203, hsa-mir-126, hsa-mir-145, hsa-mir-944, and hsa-mir-93, which have a common expression in cervical cancer and HPV. Conclusion: Overall, the gene network, including TP53, MYC, AKT1, TNF, IL6, EGFR, STAT3, CTNNB1, ESR1, and JUN, and Also, hsa-mir-145, hsa-mir-93, hsa-mir-203, and hsa-mir-126 can be regarded as a gene expression pathway in HPV-induced cervical cancer.

Identification, characterization, and anti-inflammatory activity of a lipocalin-like protein cloned from Oenanthe javanica

Identification, characterization, and anti-inflammatory activity of a lipocalin-like protein cloned from Oenanthe javanica

PE/PPE Proteome and ESX-5 Substrate Spectrum in Mycobacterium marinum.

PE/PPE proteins secreted by the ESX-5 type VII secretion system constitute a major protein repertoire in pathogenic mycobacteria and are essential for bacterial survival, pathogenicity, and host-pathogen interaction; however, little is known about their expression and secretion. The scarcity of arginine and lysine residues in PE/PPE protein sequences and the high homology of their N-terminal domains limit protein identification using classical trypsin-based proteomic methods. This study used endoproteinase AspN and trypsin to characterize the proteome of Mycobacterium marinum. Twenty-seven PE/PPE proteins were uniquely identified in AspN digests, especially PE_PGRS proteins. These treatments allowed the identification of approximately 50% of the PE/PPE pool encoded in the genome. Moreover, EspG5 pulldown assays retrieved 44 ESX-5-associated PPE proteins, covering 85% of the PPE pool in the identified proteome. The identification of PE/PE_PGRS proteins in the EspG5 interactome suggested the presence of PE-PPE pairs. The correlation analysis between protein abundance and phylogenetic relationships found potential PE/PPE pairs, indicating the presence of multiple PE/PE_PGRS partners in one PPE. We validated that EspG5 interacted with PPE31 and PPE32 and mapped critical residues for complex formation. The modified proteomic platform increases the coverage of PE/PPE proteins and elucidates the expression and localization of these proteins.

Unraveling the Mystery of Energy-Sensing Enzymes and Signaling Pathways in Tumorigenesis and Their Potential as Therapeutic Targets for Cancer.

Cancer research has advanced tremendously with the identification of causative genes, proteins, and signaling pathways. Numerous antitumor drugs have been designed and screened for cancer therapeutics; however, designing target-specific drugs for malignant cells with minimal side effects is challenging. Recently, energy-sensing- and homeostasis-associated molecules and signaling pathways playing a role in proliferation, apoptosis, autophagy, and angiogenesis have received increasing attention. Energy-metabolism-based studies have shown the contribution of energetics to cancer development, where tumor cells show increased glycolytic activity and decreased oxidative phosphorylation (the Warburg effect) in order to obtain the required additional energy for rapid division. The role of energy homeostasis in the survival of normal as well as malignant cells is critical; therefore, fuel intake and expenditure must be balanced within acceptable limits. Thus, energy-sensing enzymes detecting the disruption of glycolysis, AMP, ATP, or GTP levels are promising anticancer therapeutic targets. Here, we review the common energy mediators and energy sensors and their metabolic properties, mechanisms, and associated signaling pathways involved in carcinogenesis, and explore the possibility of identifying drugs for inhibiting the energy metabolism of tumor cells. Furthermore, to corroborate our hypothesis, we performed meta-analysis based on transcriptomic profiling to search for energy-associated biomarkers and canonical pathways.

Accurate Identification of Periplasmic Urea-binding Proteins by Structure- and Genome Context-assisted Functional Analysis

ABC transporters are ancient and ubiquitous nutrient transport systems in bacteria and play a central role in defining lifestyles. Periplasmic solute-binding proteins (SBPs) are components that deliver ligands to their translocation machinery. SBPs have diversified to bind a wide range of ligands with high specificity and affinity. However, accurate assignment of cognate ligands remains a challenging problem in SBPs. Urea metabolism plays an important role in the nitrogen cycle; anthropogenic sources account for more than half of global nitrogen fertilizer. We report identification of urea-binding proteins within a large SBP sequence family that encodes diverse functions. By combining genetic linkage between SBPs, ABC transporter components, enzymes or transcription factors, we accurately identified cognate ligands, as we verified experimentally by biophysical characterization of ligand binding and crystallographic determination of the urea complex of a thermostable urea-binding homolog. Using three-dimensional structure information, these functional assignments were extrapolated to other members in the sequence family lacking genetic linkage information, which revealed that only a fraction bind urea. Using the same combined approaches, we also inferred that other family members bind various short-chain amides, aliphatic amino acids (leucine, isoleucine, valine), γ-aminobutyrate, and as yet unknown ligands. Comparative structural analysis revealed structural adaptations that encode diversification in these SBPs. Systematic assignment of ligands to SBP sequence families is key to understanding bacterial lifestyles, and also provides a rich source of biosensors for clinical and environmental analysis, such as the thermostable urea-binding protein identified here.

From Sequence to Solution: Intelligent Learning Engine Optimization in Drug Discovery and Protein Analysis

This study introduces the intelligent learning engine (ILE) optimization technology, a novel approach designed to revolutionize screening processes in bioinformatics, cheminformatics, and a range of other scientific fields. By focusing on the efficient and precise identification of candidates with desirable characteristics, the ILE technology marks a significant leap forward in addressing the complexities of candidate selection in drug discovery, protein classification, and beyond. The study’s primary objective is to address the challenges associated with optimizing screening processes to efficiently select candidates across various fields, including drug discovery and protein classification. The methodology employed involves a detailed algorithmic process that includes dataset preparation, encoding of protein sequences, sensor nucleation, and optimization, culminating in the empirical evaluation of molecular activity indexing, homology-based modeling, and classification of proteins such as G-protein-coupled receptors. This process showcases the method’s success in multiple sequence alignment, protein identification, and classification. Key results demonstrate the ILE’s superior accuracy in protein classification and virtual high-throughput screening, with a notable breakthrough in drug development for assessing drug-induced long QT syndrome risks through hERG potassium channel interaction analysis. The technology showcased exceptional results in the formulation and evaluation of novel cancer drug candidates, highlighting its potential for significant advancements in pharmaceutical innovations. The findings underline the ILE optimization technology as a transformative tool in screening processes due to its proven effectiveness and broad applicability across various domains. This breakthrough contributes substantially to the fields of systems optimization and holds promise for diverse applications, enhancing the process of selecting candidate molecules with target properties and advancing drug discovery, protein classification, and modeling.

Analysis of expression characteristics and identification of interaction proteins of transcription factor BnaABI5 in Brassica napus.

Rapeseed is one important oil crop in China. However, its planting benefit is frequently affected by environmental stresses such as drought in the northwest region of China. The abscisic acid(ABA) signaling pathway plays an important role in plant abiotic stress response and tolerance, and ABFs/AREBs(ABA-responsive element binding factors/ABA-responsive element binding proteins) are the core transcription factors that regulate the expression of ABA-responsive genes. To dissect the key transcription factors mediated abiotic stress, we mainly characterized abscisic acid insensitive 5(BnaABI5) in rapeseed, including its subcellular localization, expression pattern in response to various stress and tissue-specific expression analysis, transcriptional activity analysis as well as interaction screening with BnaMPKs(mitogen-activated protein kinases). Our results showed that the BnaABI5-GFP fusion protein was localized in the nucleus, and its transcript level is induced by drought stress and was mainly expressed in the roots of rapeseed. Furthermore, BnaABI5 showed transcriptional activation activity through a yeast transactivation assay and it also activated the promoter activity of EM6 target gene in the transient expression system in tobacco leaves. Moreover, BnaABI5 interacted with BnaMPK6 and BnaMPK13 through BiFC and Y2H analysis. This study preliminarily explored the expression characteristics of transcription factor BnaABI5 and its interaction with BnaMPKs, which might help us for further understanding the function of BnaABI5.

GraphNABP: Identifying nucleic acid-binding proteins with protein graphs and protein language models

The computational identification of nucleic acid-binding proteins (NABP) is of great significance for understanding the mechanisms of these biological activities and drug discovery. Although a bunch of sequence-based methods have been proposed to predict NABP and achieved promising performance, the structure information is often overlooked. On the other hand, the power of popular protein language models (pLM) has seldom been harnessed for predicting NABPs. In this study, we propose a novel framework called GraphNABP, to predict NABP by integrating sequence and predicted 3D structure information. Specifically, sequence embeddings and protein molecular graphs were first obtained from ProtT5 protein language model and predicted 3D structures, respectively. Then, graph attention (GAT) and bidirectional long short-term memory (BiLSTM) neural networks were used to enhance feature representations. Finally, a fully connected layer is used to predict NABPs. To the best of our knowledge, this is the first time to integrate AlphaFold and protein language models for the prediction of NABPs. The performances on multiple independent test sets indicate that GraphNABP outperforms other state-of-the-art methods. Our results demonstrate the effectiveness of pLM embeddings and structural information for NABP prediction. The codes and data used in this study are available at https://github.com/lixiangli01/GraphNABP.

Identification of antigenic proteins from the venom of Malaysian snakes using immunoprecipitation assay and tandem mass spectrometry (LC-MS/MS)

Snake envenomation poses a significant risk to Malaysians and country visitors. Malaysia witnesses an estimated 650 snake bites per 100,000 population annually. The primary treatment for snake envenomation involves administering antivenom derived from horses, despite its drawbacks, such as anaphylactic reactions and serum sickness. Identifying the venom proteome is crucial for understanding and predicting the clinical implications of envenomation and developing effective treatments targeting specific venom proteins. In this study, we employ an immunoprecipitation assay followed by LC-MS/MS to identify antigenic proteins in five common venomous snakes in Malaysia compassing of two families which are pit vipers, (Calloselasma rhodostoma and Cryptelytrops purpureomaculatus) and cobras (Ophiophagus hannah, Naja kaouthia, and Naja sumatrana). The immunoprecipitation assay utilises a 2% agarose gel, allowing antigenic proteins to diffuse and bind with antibodies in the antivenom. The antivenom utilised in this research was procured from the Queen Saovabha Memorial Institute (QSMI), Thailand, including king cobra antivenom (KCAV), cobra antivenom (CAV), Malayan pit viper antivenom (MPAV), Russell's viper antivenom (RPAV), hematopolyvalent antivenom (HPAV), neuropolyvalent antivenom (NPAV), banded krait antivenom (BKAV), and Malayan krait antivenom (MKAV). The protein identified through these interactions which are exclusive to the cobras are three-finger toxins (3FTXs) while snake C-type lectins (Snaclecs) are unique to the pit vipers. Common protein that are present in both families are L-amino acid oxidase (LAAO), Phospholipase A2 (PLA2), and snake venom metalloproteinase (SVMP). Identifying these proteins is vital for formulating a broad-spectrum antivenom applicable across multiple species.

Identification of Key Proteins Related to Cashmere Fiber Diameter by Integrated Proteomics and Bioinformatic Analyses in the Alpas and Alxa Goat Breeds

Background: Goats (Capra hircus) have always been a source of fiber for human use and hold an important place in international high-end textiles. Fiber diameter is the most concerning economic indicator for producers. Understanding the formation mechanism of fiber diameter and its related key proteins can help optimize and control the production of cashmere. Methods: Cashmere goats (n = 36) of the Alpas (n = 18) and Alxa (n = 18) breeds, with a similar age (2 years old) and live weight (25–26 kg), were selected from the Yiwei White Cashmere Goat Breeding Farm, Erdos, Inner Mongolia. Using phenotypic indicators, we evaluated the diameter of the cashmere fibers in Alxa and Alpas goats. We also used electron microscopy to examine the cashmere fiber’s structure and label-free liquid chromatography–tandem mass spectrometry to determine the protein content of the two cashmere fibers. The proteins affecting fiber diameter were identified and analyzed by Western blot, Co-Immunoprecipitation, and bioinformatics analysis. Results: The average diameter of the Alxa breed was smaller (p < 0.05) than that of the Alpas breed (Alxa’s cashmere vs. Alpas’ cashmere). Proteomics technology enabled the highly confident detection of 171 proteins. A total of 68 differentially expressed proteins were identified in the two types of cashmere; 131 proteins were specifically expressed in Alpas goats, and 40 proteins were specifically expressed in Alxa goats. A key protein group that could cause variations in fiber diameter was found using the protein–protein interaction network. To ascertain the reason for the variation in fiber diameter, a structural study of the major protein groups was carried out. Conclusions: KRT10, KRT14, KRT17, and KRT82 are the main proteins impacting the diameter difference, and they have a substantial effect on the average fiber diameter.