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PubMed1992

Microsequences of 145 proteins recorded in the two-dimensional gel protein database of normal human epidermal keratinocytes.

Microsequencing of proteins recovered from two-dimensional (2-D) gels is being used systematically to identify proteins in the master human keratinocyte 2-D gel database. To date, about 250 protein spots recorded in human 2-D gel databases have been microsequenced and, of these, 145 are recorded in the keratinocyte database under the entry partial amino acid sequence. Coomassie Brilliant Blue-stained protein spots cut from several (up to 40) dry gels were concentrated by elution-concentration gel electrophoresis, electroblotted onto PVDF membranes and digested in situ with trypsin. Eluting peptides were separated by reversed-phase HPLC, collected individually and sequenced. Computer search using the FASTA and TFASTA programs from Genetics Computer Group indicated that 110 of the microsequenced polypeptides shared significant similarity with proteins contained in the PIR, Mipsx or GenEMBL databases. Only 35 polypeptides corresponded to hitherto unknown proteins. Peptide sequences of all 145 proteins are listed together with their coordinates (apparent molecular weight and pI) in the keratinocyte database.

Amino Acid Sequence↗

PubMed1992

Fast structure alignment for protein databank searching.

A fast method is described for searching and analyzing the protein structure databank. It uses secondary structure followed by residue matching to compare protein structures and is developed from a previous structural alignment method based on dynamic programming. Linear representations of secondary structures are derived and their features compared to identify equivalent elements in two proteins. The secondary structure alignment then constrains the residue alignment, which compares only residues within aligned secondary structures and with similar buried areas and torsional angles. The initial secondary structure alignment improves accuracy and provides a means of filtering out unrelated proteins before the slower residue alignment stage. It is possible to search or sort the protein structure databank very quickly using just secondary structure comparisons. A search through 720 structures with a probe protein of 10 secondary structures required 1.7 CPU hours on a Sun 4/280. Alternatively, combined secondary structure and residue alignments, with a cutoff on the secondary structure score to remove pairs of unrelated proteins from further analysis, took 10.1 CPU hours. The method was applied in searches on different classes of proteins and to cluster a subset of the databank into structurally related groups. Relationships were consistent with known families of protein structure.

Amino Acid Sequence↗

PubMed2025

Extensive Analysis of Genetic Diversity in HLA-DMA, HLA-DMB, HLA-DOA and HLA-DOB: Characterisation of 236 Novel Alleles.

HLA-DMA, -DMB, -DOA and -DOB are non-classical HLA Class II genes that play a crucial role in the selection of highly stable HLA Class II/peptide complexes on antigen-presenting cells. Although the genes were initially thought to have a limited diversity with less than 13 alleles per gene documented in the IPD-IMGT/HLA Database in 2022, recent studies suggest a potential impact of certain alleles on the outcome of hematopoietic cell transplantation. To gain a deeper understanding of allelic diversity, we sequenced HLA-DMA, -DMB, -DOA and -DOB of 1880 potential stem cell donors from Germany, Poland, Great Britain and Chile, achieving full-gene resolution. Remarkably, we identified 3968 previously undescribed sequences, including 28 distinct novel proteins. The observed allele frequencies were consistent across all studied populations with one dominating protein for each gene: HLA-DMA*01:01 (> 77%), HLA-DMB*01:01 (> 63%), HLA-DOA*01:01 (> 97%) and HLA-DOB*01:01 (> 77%). Notably, a much higher diversity was observed in full-genomic resolution. Finally, we submitted 51 distinct novel sequences for HLA-DMA, 58 for HLA-DMB, 80 for HLA-DOA and 47 for HLA-DOB to the IPD-IMGT/HLA Database. This comprehensive reference database update will not only simplify future genotyping of HLA-DMA, -DMB, -DOA and -DOB but will hopefully also enhance our understanding of the complex process of peptide selection and loading to the HLA Class II proteins.

Humans↗

PubMed2026

High-Resolution Chromosome-Level Genome Assembly and Annotation of Triplophysa stewarti, an Endemic Plateau Loach from the Qinghai-Tibet Plateau.

The bottom-dwelling fish Triplophysa stewarti, endemic to the Qinghai-Tibet Plateau, is a valuable model for studying high-altitude adaptation in aquatic ecosystems. However, the lack of a high-quality reference genome has hindered comparative genomic and evolutionary studies within this genus. Here, we present a chromosome-level genome assembly for T. stewarti, generated using PacBio HiFi long-read sequencing and Hi-C scaffolding. The 697.9 Mb assembly is highly continuous (scaffold N50 of 253.58 Mb) and encompasses 25 chromosomes, representing 92.65% of the genome. BUSCO analysis indicated a 98.4% completeness, supporting the high quality of the assembly. We annotated 28,009 protein-coding genes, with 97.04% being functionally assigned across multiple databases (NR, UniProt, KEGG, GO, Pfam and InterPro). Additionally, repetitive elements constituted 42.47% of the genome, and we identified 52,709 non-coding RNAs. This high-quality reference genome provides a fundamental resource for exploring the adaptive evolution, population structure, and conservation genetics of T. stewarti and related species on the Qinghai-Tibet Plateau.

Animals↗

PubMed1992

Human cerebrospinal fluid protein database: edition 1992.

Two-dimensional electrophoresis maps of human cerebrospinal fluid proteins are presented in the form of labeled images. 931 protein spots are identified in spinal fluid from a normal volunteer. Distinct spots that represent variants of the same protein, especially posttranslational modifications, are estimated to reduce the 931 different spots to < 200 different proteins. 248 spots of 29 protein groups have been identified and are indicated on enlargements of specific gel regions. The distribution of protein abundance, mass, charge and shape characteristics of these normal 931 spinal fluid spots are graphically profiled. Analysis of the shape parameter "vertical height: width ratio" reveals that a ratio > 3.5 correlates with glycoproteins, enabling their identification simply by image analysis. Proteins that are not present on the normal map, but appear in spinal fluid in patients with schizophrenia and Creutzfeldt-Jakob disease are illustrated on additional maps.

Cerebrospinal Fluid Proteins↗

PubMed1992

Environment-specific amino acid substitution tables: tertiary templates and prediction of protein folds.

The local environment of an amino acid in a folded protein determines the acceptability of mutations at that position. In order to characterize and quantify these structural constraints, we have made a comparative analysis of families of homologous proteins. Residues in each structure are classified according to amino acid type, secondary structure, accessibility of the side chain, and existence of hydrogen bonds from the side chains. Analysis of the pattern of observed substitutions as a function of local environment shows that there are distinct patterns, especially for buried polar residues. The substitution data tables are available on diskette with Protein Science. Given the fold of a protein, one is able to predict sequences compatible with the fold (profiles or templates) and potentially to discriminate between a correctly folded and misfolded protein. Conversely, analysis of residue variation across a family of aligned sequences in terms of substitution profiles can allow prediction of secondary structure or tertiary environment.

Amino Acid Sequence↗

PubMed2026

FANTASIA suite: a reproducible and configurable framework for embedding-based functional annotation of proteins.

Embedding-based annotation transfer is increasingly used for protein function inference due to protein language models capture sequence, structural, and functional signals that may extend beyond conventional pairwise similarity. However, systematic application of these approaches requires control over model choice, reference composition, lookup parameters, evidence traceability, and output formats. We developed the FANTASIA suite, a configurable framework for embedding-based functional annotation of proteins. The suite combines a database-backed implementation for reproducible and extensible analyses with a portable flat-file implementation for rapid local annotation and pipeline integration. Using non-model and model-organism proteomes, we show that larger neighbourhood sizes remain practical for proteome-scale analyses and that taxonomy and sequence-identity filtering support leakage-aware benchmarking. We also compare the supported models with baseline methods through external CAFA5 evaluation and provide practical guidance based on empirical evidence variables. FANTASIA provides a controlled, scalable, and reproducible framework for extending functional annotation across the rapidly expanding diversity of sequenced organisms.

Software↗

PubMed2025

Network pharmacology and molecular docking to explore the active compounds and mechanisms of Jerusalem artichoke for treating diabetes.

The effective components and mechanism of Jerusalem artichokes (JAs) in lowering blood glucose were studied through network pharmacology and molecular docking. The active compounds of Jerusalem artichoke were obtained by referring to the literature, and the active compounds were screened. The targets were predicted by the SwissTargetPrediction database, and the disease targets were screened using GeneCard, Disgenet, and OMIM databases. The protein-protein interaction (PPI) network diagram was constructed using the STRING database, and the intersection target was analyzed by gene ontology (GO) biological function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses using the David database. Finally, molecular docking was verified using AutoDockTools1.5.7 software. After screening, 412 gene targets, 476 disease targets, and 64 intersection targets were identified. The results of GO biological function analysis and KEGG pathway analysis showed that the technology was involved in multiple biological processes and regulatory pathways for hypoglycemia, such as the HIF-1, PI3K-Akt, and AMPK signaling pathways. Molecular docking results showed that Jasmonate, Liquiritigenin and Inulin of JAs had strong binding effects with PPARG and STAT3. JAs exert hypoglycemic effects through multi-component, multi-target and multi-pathway. In summary, this study investigated the hypoglycemic mechanism of JAs using network pharmacology and molecular interconnection technology, and concluded that JAs exert hypoglycemic effects through multiple components, targets, and pathways, which provides a theoretical basis for the study of JAs.

Molecular Docking Simulation↗

PubMed2026

In Situ Hybridization and RT-PCR Detection of Nervous Necrosis Virus in Fourfinger Threadfin, Eleutheronema tetradactylum, in Taiwan.

Between April and July 2020, suspected outbreaks of nervous necrosis virus (NNV) infection were observed in fourfinger threadfin (Eleutheronema tetradactylum) fingerling hatcheries in Pingtung County, southern Taiwan. Affected fish exhibited spiral swimming behaviour and abdominal distension, resulting in mortality rates between 50% and 100%. Histopathological examination showed severe vacuolation in the brain and ocular tissues, with large oval and/or irregular basophilic cytoplasmic inclusion bodies in the brain. Phylogenetic analysis of the viral replicase (RNA1) and capsid protein (RNA2) genes revealed high nucleotide sequence identities among the isolates in this study, with sequence similarity rates of 96.9%-99% for RNA1 and 98.2%-99.0% for RNA2 compared to RGNNV reference strains available in the NCBI GenBank database. This is the first detection of betanodavirus in fourfinger threadfin in Taiwan, using RT-PCR and ISH. A positive correlation between elevated water temperatures and disease severity indicates the need for year-round surveillance and genomic analysis to clarify the epidemiology of FTNNV. The data suggest that infected eggs may facilitate the vertical transmission of Betanodavirus. Crucially, utilising virus-free broodstock, alongside routine health screening and environmental control, is essential for mitigating NNV risks in fourfinger threadfin aquaculture.

Animals↗

PubMed2025

PEELing: an integrated and user-centric platform for spatially resolved proteomics data analysis.

SUMMARY: Molecular compartmentalization is vital for cellular physiology. Spatially resolved proteomics allows biologists to survey protein composition and dynamics with subcellular resolution. Here, we present PEELing, an integrated package and user-friendly web service for analyzing spatially resolved proteomics data. PEELing assesses data quality using curated or user-defined references, performs cutoff analysis to remove contaminants, connects to databases for functional annotation, and generates data visualizations-providing a streamlined and reproducible workflow to explore spatially resolved proteomics data. AVAILABILITY AND IMPLEMENTATION: PEELing and its tutorial are publicly available at https://peeling.janelia.org/ (Zenodo DOI: 10.5281/zenodo.15692517). A Python package of PEELing is available at https://github.com/JaneliaSciComp/peeling/ (Zenodo DOI: 10.5281/zenodo.15692434).

Proteomics↗

PubMed2025

HI-FEVER: a Nextflow pipeline for the high-throughput discovery and annotation of endogenous viral elements.

SUMMARY: Endogenous viral elements (EVEs) offer valuable insights into virus and host evolution, but their detection remains computationally and biologically challenging. We present HI-FEVER, a user-friendly Nextflow pipeline for the discovery of EVEs in eukaryotic host genomes. HI-FEVER is highly parallelizable and customizable, ensuring computational efficiency while allowing researchers to fine-tune parameters to their specific needs. Its output provides a comprehensive analysis of discovered EVEs, including detailed annotations which can provide evolutionary insights. HI-FEVER scales seamlessly to handle millions of viral protein queries across multiple host genomes on both laptops and high-performance computing nodes. AVAILABILITY AND IMPLEMENTATION: The HI-FEVER source code is available on GitHub at https://github.com/PaleovirologyLab/hi-fever. Minimal reference databases, test datasets and benchmarking results are hosted on the Open Science Framework at https://osf.io/y357r. A detailed wiki is available at https://github.com/PaleovirologyLab/hi-fever/wiki, including usage instructions, parameter descriptions, and guidance on interpreting outputs. The pipeline includes a Pixi environment compatible with Conda and Apptainer containerization, and Docker images. HI-FEVER has been tested on Linux, Windows (via WSL2), and macOS (Intel and ARM64).

Software↗

PubMed2026

The Role of Small Segmental Duplications in Generating Identical Isoforms Through Alternative Splicing Sites.

Alternative splicing plays a crucial role in expanding proteomic diversity but can also generate identical isoforms under certain conditions. While mutually exclusive splicing of tandem exons has occasionally been reported to produce identical isoforms, the extent to which other splicing events contribute to this phenomenon remains unclear. In this study, we demonstrate that alternative 5' and 3' splice site selection can also lead to the formation of identical isoforms, providing an additional type of splicing event for functional redundancy in transcriptomes. To address this, we analyzed reference genome annotations from 15 plant species, including Arabidopsis thaliana and wheat (Triticum aestivum), obtained from the RefSeq database. Identical isoforms were computationally defined as transcripts with distinct exon-intron structures but identical coding sequences. Our analysis reveals that the majority of alternative 5' and 3' fragments originate from small segmental duplications, suggesting that sequence repetition within gene regions facilitates the emergence of such splicing patterns. We also observed differences in the annotated 5' UTRs of some identical isoforms. However, since the alternative splicing sites themselves were not located within UTRs, these differences may reflect annotation uncertainty rather than genuine AS-derived variation. Given that UTR predictions in reference databases are not always precise, such observations should be interpreted cautiously. Expression analysis using an isoform-specific k-mer approach confirmed that identical isoforms can be differentially regulated. These findings suggest that, beyond expanding protein diversity, alternative splicing can also generate redundant isoforms that are differentially expressed at the RNA level, indicating potential regulatory roles. By elucidating the structural and regulatory factors contributing to the formation and retention of identical isoforms, our study provides new insights into the evolutionary and functional significance of alternative splicing in plants.

Alternative Splicing↗

PubMed2025

In silico prediction of the impact of genomic variations in the small conductance calcium activated potassium channel SK3 structure and function.

The small-conductance calcium-activated potassium channel SK3, encoded by the KCNN3 gene, plays a critical role in regulating dopaminergic neuron (DN) firing patterns by modulating after hyperpolarization currents. SK3 dysfunction has been implicated in neuropsychiatric and neurodegenerative disorders. We analyzed structural and functional consequences of KCNN3 splicing and genetic variation. Alternative splicing variants of the KCNN3 gene were retrieved from the Ensembl database and aligned using T-Coffee, manually inspected and curated. Protein domains were identified with Pfam 35.0, SMART 9.0, and InterPro 98.0, and visualized. An AlphaFold2 model of SK3 full-length protein (UniProt: Q9UGI6) used as reference and structural models of its splicing variants were predicted with ColabFold. Functional domains (S1-S6 transmembrane helices, H5 pore loop, and calmodulin-binding) were defined and superimposed onto the AlphaFold2 reference. Domain integrity was assessed based on completeness of all expected residue indices within each functional region. SNPs and CNVs across all coding KCNN3 splicing variants were analyzed, classified, and filtered to isolate pathogenic variants prioritizing non-synonymous amino acid substitutions. Differential variant impacts across splicing isoforms were assessed by mapping variant positions to individual transcript protein sequences and used to predict functional consequences. Two long and two short splicing variants are known. Short variants lack the motif required for potassium channels. Pathogenic variants result from missense mutations resulting in amino acid substitutions. In all cases, the consequential effects depend on the specific location and role of the amino acid being changed.

SK3 channels↗

PubMed2026

A gap-free, telomere-to-telomere chromosome-scale genome assembly of the mangrove red snapper, Lutjanus argentimaculatus.

The mangrove red snapper (Lutjanus argentimaculatus) is a commercially important marine fish species in the Indo-Pacific region. Despite its significant economic value for aquaculture, existing genomic resources remain fragmented, limiting the advancement of molecular breeding and functional genomic studies. Here, we present a gap-free, telomere-to-telomere (T2T) genome assembly of L. argentimaculatus, generated using a hybrid approach combining PacBio HiFi, Oxford Nanopore ultra-long reads and Hi-C technology. The resulting assembly comprises exactly 24 scaffolds spanning 1.03 Gb, perfectly matching the haploid chromosome number with a contig N50 of 46.17 Mb. Notably, this assembly resolves all physical gaps present in previous versions, achieving a BUSCO completeness score of 98.2%. Comprehensive genome annotation successfully predicted 23,167 protein-coding genes. Among these, 22,067 genes (95.25%) were functionally annotated across major public databases, including eggNOG, InterPro, and Swiss-Prot. Furthermore, structural analysis successfully identified 19 telomeres and 20 centromeres, validating the chromosomal integrity. This high-fidelity, gap-free reference genome provides a robust foundation for comparative genomics, population genetics, and the genetic improvement of Lutjanidae species.

Animals↗

PubMed2025

mettannotator: a comprehensive and scalable Nextflow annotation pipeline for prokaryotic assemblies.

SUMMARY: In recent years, there has been a surge in prokaryotic genome assemblies, coming from both isolated organisms and environmental samples. These assemblies often include novel species that are poorly represented in reference databases creating a need for a tool that can annotate both well-described and novel taxa, and can run at scale. Here, we present mettannotator-a comprehensive, scalable Nextflow pipeline for prokaryotic genome annotation that identifies coding and noncoding regions, predicts protein functions, including antimicrobial resistance, and delineates gene clusters. The pipeline summarizes these results in a GFF (General Feature Format) file that can be easily utilized in downstream analysis or visualized using common genome browsers. Here, we show how it works on 200 genomes from 29 prokaryotic phyla, including isolate genomes and known and novel metagenome-assembled genomes, and present metrics on its performance in comparison to other tools. AVAILABILITY AND IMPLEMENTATION: The pipeline is written in Nextflow and Python and published under an open source Apache 2.0 licence. Instructions and source code can be accessed at https://github.com/EBI-Metagenomics/mettannotator. The pipeline is also available on WorkflowHub: https://workflowhub.eu/workflows/1069.

Software↗

PubMed1990

Flexible protein sequence patterns. A sensitive method to detect weak structural similarities.

The concept of a flexible protein sequence pattern is defined. In contrast to conventional pattern matching, template or sequence alignment methods, flexible patterns allow residue patterns typical of a complete protein fold to be developed in terms of residue positions (elements), separated by gaps of defined range. An efficient dynamic programming algorithm is presented to enable the best alignment(s) of a pattern with a sequence to be identified. The flexible pattern method is evaluated in detail by reference to the globin protein family, and by comparison to alignment techniques that exploit single sequence, multiple sequence and secondary structural information. A flexible pattern derived from seven globins aligned on structural criteria successfully discriminates all 345 globins from non-globins in the Protein Identification Resource database. Furthermore, a pattern that uses helical regions from just human alpha-haemoglobin identified 337 globins compared to 318 for the best non-pattern global alignment method. Patterns derived from successively fewer, yet more highly conserved positions in a structural alignment of seven globins show that as few as 38 residue positions (25 buried hydrophobic, 4 exposed and 9 others) may be used to uniquely identify the globin fold. The study suggests that flexible patterns gain discriminating power both by discarding regions known to vary within the protein family, and by defining gaps within specific ranges. Flexible patterns therefore provide a convenient and powerful bridge between regular expression pattern matching techniques and more conventional local and global sequence comparison algorithms.

Amino Acid Sequence↗

PubMed1991

A two-dimensional gel database of rat liver proteins useful in gene regulation and drug effects studies.

A standard two-dimensional (2-D) protein map of Fischer 344 rat liver (F344MST3) is presented, with a tabular listing of more than 1200 protein species. Sodium dodecyl sulfate (SDS) molecular mass and isoelectric point have been established, based on positions of numerous internal standards. This map has been used to connect and compare hundreds of 2-D gels of rat liver samples from a variety of studies, and forms the nucleus of an expanding database describing rat liver proteins and their regulation by various drugs and toxic agents. An example of such a study, involving regulation of cholesterol synthesis by cholesterol-lowering drugs and a high-cholesterol diet, is presented. Since the map has been obtained with a widely used and highly reproducible 2-D gel system (the Iso-Dalt system), it can be directly related to an expanding body of work in other laboratories.

Animals↗

PubMed1992

Comparison of six microcomputer dietary analysis systems with the USDA Nutrient Data Base for Standard Reference.

We compared the general operating features and nutrient databases of six microcomputer dietary analysis systems. A 3-day food record with 73 food items was entered into each program; nutrient averages were compared with the US Department of Agriculture Nutrient Data Base for Standard Reference (USDA NDB), full version, release 9, for microcomputers. The six programs were found to vary widely in cost, number of foods and nutrients in the database, use of non-USDA data and imputation of data for missing values, number of print/export options, time to analyze the 3-day food record, and overall ease of use. Although all of the microcomputer dietary analysis systems were within 7% of the USDA NDB for energy, protein, total fat, and total carbohydrates, the proportion of other nutrients varying more than 15% from the USDA NDB varied considerably between programs. Variance among programs for 3-day food record nutrient values occurred because of differences in the number of food items included in the database (leading to varying degrees of substitution), the recency of the nutrient data (whether or not the most recent USDA releases had been incorporated), and the number of missing values (the degree to which non-USDA sources or estimated calculations were used to fill in the blanks from the USDA standard). Our results demonstrate that it is important for each dietitian to carefully choose a microcomputer dietary analysis system that is suitable to specific and predetermined needs.

Databases, Factual↗