The second half of 2025 has been an exceptionally productive period for the Istanbul Technical University Department of Molecular Biology and Genetics, marked by a diverse range of high-impact publications and international collaborations. From cutting-edge genomic analyses and computational vaccine design to metagenomic exploration of Türkiye’s extreme ecosystems and advances in proteomics and enzyme engineering, our faculty and researchers have contributed significantly to globally relevant scientific questions. These achievements, published across Q1 and Q2 journals, reflect the department’s growing momentum in interdisciplinary research and its commitment to generating knowledge with both fundamental and applied impact. As Istanbul Technical University's Department of Molecular Biology and Genetics continues to expand its scientific footprint, the department stands out as a dynamic hub of innovation, academic excellence, and international engagement.




1. CRISPR-Cas repertoire of Kluyvera ascorbata

In the article titled “The CRISPR-Cas Repertoire of Kluyvera ascorbata: Insights from Genomic Data, published in Current Genetics“(Q2 in Scopus ranking), Assis. Prof. Dr. Çiğdem Yılmaz Çolak presents the first comprehensive investigation of adaptive immunity elements in this clinically relevant species. Through comparative genome mining across the Kluyvera genus, the study identifies K. ascorbata as the only member carrying a complete and structurally coherent Type I-E clustered regularly interspaced short palindromic repeats (CRISPR)-Cas system. The work highlights the evolutionary traces of past interactions with phages and plasmids and points to the potential functional significance of these immune components in genome defense. These findings provide an important framework for future research exploring the diagnostic, evolutionary, and mechanistic implications of CRISPR-Cas elements in Kluyvera ascorbata.

Kluyvera species have recently gained attention as opportunistic pathogens and reservoirs of antibiotic-resistance genes, yet their adaptive immune systems remain poorly described. By examining 13 publicly available genomes, this study provides the first focused characterization of CRISPR-Cas elements in the genus and reveals that only Kluyvera ascorbata carries a complete, identifiable CRISPR-Cas system, specifically a Type I-E architecture containing core genes and multiple CRISPR arrays with conserved repeat motifs. Spacer analysis identified matches to phage and plasmid sequences, indicating historical interactions with mobile genetic elements and suggesting a potentially functional immune repertoire that may influence horizontal gene transfer dynamics. Phylogenetic analysis placed the K. ascorbata Cas3 protein close to those of other Enterobacteriaceae, including Salmonella and Citrobacter, pointing to shared evolutionary origins or lateral acquisition. The absence or fragmentation of CRISPR-Cas systems in other Kluyvera genomes raises questions regarding selective pressures shaping system retention or loss within the genus.

The findings offer a basis for exploring CRISPR-mediated phage resistance, genome stability, and epidemiological applications, while also suggesting that CRISPR-Cas markers could support improved species-level identification in clinically relevant Kluyvera isolates.

DOI: https://doi.org/10.1007/s00294-025-01332-x

2. Multi-Epitope Vaccine for Piscirickettsia salmonis

In the article titled “Targeting potential virulence factors of Piscirickettsia salmonis for vaccine development through an immunoinformatics approach” published in Aquaculture International (Q2 in Scopus ranking), Assist. Prof. Dr. Çiğdem Yılmaz Çolak presents a computational framework for developing a next-generation vaccine candidate against the aquaculture pathogen P. salmonis. The study identifies four virulence-associated proteins as potential antigen sources and employs a reverse-vaccinology strategy to pinpoint immunogenic epitopes relevant for Atlantic salmon. These epitopes were integrated into a multi-epitope subunit construct whose structural properties and receptor-binding potential were extensively evaluated through in silico modelling. The findings suggest that the designed construct exhibits promising stability and immune-stimulatory potential, emphasizing the value of immunoinformatics in accelerating antigen discovery for aquaculture vaccines and laying the groundwork for future experimental validation.

The Gram-negative bacterium Piscirickettsia salmonis is the etiologic agent of piscirickettsiosis, which significantly impacts salmon aquaculture worldwide. Although commercial vaccines exist, their efficacy remains sub-optimal, motivating the search for more effective antigenic strategies. In the current study, authors selected four putative virulence-associated outer-membrane or secretion-system-related proteins (TolC, BamB, LptD, and OmpA) as vaccine antigen targets. Through a reverse-vaccinology and immunoinformatics pipeline, linear B-cell epitopes and T-cell epitopes were predicted from these proteins and evaluated for antigenicity, allergenicity, toxicity and conservation. Selected epitopes were stitched into a chimeric multi-epitope construct employing linkers to facilitate proper folding and protease cleavage. The construct’s physicochemical parameters (such as molecular weight, pI, instability, aliphatic index) were computed using ProtParam. Its tertiary structure was predicted using AlphaFold, then refined with GalaxyRefine and validated with ProSA and Ramachandran plot analysis. Next, molecular docking simulations between the vaccine construct and Atlantic salmon TLR 5 receptor were performed, revealing a stable binding conformation. Normal mode analysis and CABS-flex simulations assessed structural flexibility, root mean-square fluctuation, covariance and deformability, confirming dynamic stability. Immune-response simulations indicated favorable cytokine profiles and B/T-cell activation.

The study concludes that the designed multi-epitope vaccine holds promising in silico performance and represents a candidate for further experimental validation in fish. The work aligns with the growing interest in next-generation subunit vaccines for aquaculture pathogens and leverages advanced computational modelling to streamline antigen design.

DOI: https://doi.org/10.1007/s10499-025-02334-9

3.  Exploring Türkiye’s Extreme P450 Diversity

A study titled “Exploring Extreme Environments in Türkiye for Novel P450s Through Metagenomic Analysis” conducted by Research Assistant Hande Mumcu as part of her PhD thesis under the supervision of Prof. Dr. Nevin Gül Karagüler and in collaboration with researchers from The University of Queensland, has been published in the peer-reviewed journal PLOS ONE (Q1 in Scopus ranking). By applying high-throughput metagenomic strategies, the team investigated Türkiye’s diverse extreme habitats and uncovered a rich, previously uncharacterized repertoire of microbial cytochrome P450 enzymes. Their findings highlight how distinct environmental conditions shape oxidative metabolism, positioning these ecosystems as valuable sources of novel biocatalysts with strong industrial potential.

Cytochrome P450 enzymes (P450s) are among the most versatile oxidative biocatalysts known, catalyzing highly regio- and stereoselective reactions with pronounced substrate specificity. When these enzymes originate from extremophilic microorganisms, they often combine this catalytic precision with an ability to function under high salinity, temperature, or acidity, which makes them particularly attractive for applied biocatalysis. However, much of this extremophile P450 diversity remains unexplored. In this study, researchers from the Department of Molecular Biology and Genetics at Istanbul Technical University, in collaboration with researchers from the University of Queensland and the University of Tennessee Health Science Center, investigated six geochemically distinct extreme environments across Türkiye. The sites comprised three hypersaline systems (Lake Acıgöl, Tuz Gölü, and Gömeç), two hydrothermal springs (Hisaralan and Armutlu), and an acid mine drainage site (Balya). Sediment, water, or salt-crystal samples were collected depending on the local matrix, and environmental DNA was extracted for high-throughput shotgun metagenomic sequencing.

 Quality-controlled reads from each site were assembled and then processed through a comprehensive binning pipeline to reconstruct metagenome-assembled genomes (MAGs). This workflow yielded 1,138 non-redundant bins, of which 171 met specific quality thresholds and collectively represented four archaeal and 28 bacterial phyla, including many lineages with no previously available genomes. These MAGs provided the taxonomic framework required to interpret enzyme diversity within and across sites. Putative P450s were identified by screening the predicted protein sets against profile hidden Markov models and curated reference databases, followed by stringent filtering for full-length sequences containing the canonical heme-binding motif. A total of 311 high-confidence microbial P450s were discovered. These enzymes were classified into 87 families and 158 subfamilies, including eight novel families and 49 new subfamilies that expand the current cytochrome P450 nomenclature. A large subset of the sequences could be linked back to specific MAGs, anchoring P450 diversity in well-defined microbial hosts. The six sampling sites exhibited distinct P450 “signatures” that reflected their environmental conditions. Thermophilic P450s were enriched in the hydrothermal systems Hisaralan and Armutlu, acidophilic lineages dominated the Balya mine drainage, and halophilic P450s prevailed in the hypersaline lakes. The CYP107 family emerged as a recurrent lineage shared by nearly all sites, while families such as CYP174, CYP197, CYP1103, and CYP108 were strongly site-enriched, underscoring local adaptation of oxidative metabolism to geochemical context.

Overall, the work demonstrates that Türkiye’s extreme habitats host a dense and taxonomically diverse reservoir of cytochrome P450 enzymes. By combining deep shotgun metagenomics with advanced binning and rigorous functional annotation, this study establishes a rich resource of candidate extremozymes for future biochemical, structural, and biotechnological characterisation, with potential applications in industrial biocatalysis and environmental remediation. For İTÜ and its partners, these findings reveal how national biodiversity, combined with cutting-edge metagenomics, can guide enzyme discovery efforts with global impact.

DOI: https://doi.org/10.1371/journal.pone.0330523

4. Prof. Dr. Gizem Dinler Doğanay Contributes to Landmark Study Mapping the Human Pan-Disease Blood Proteome

A major international study co-authored by Prof. Dr. Gizem Dinler Doğanay of Istanbul Technical University’s Department of Molecular Biology and Genetics has produced one of the most comprehensive characterizations of the circulating human blood proteome to date. Published in Science, the work integrates proteomic measurements from 8,262 individuals—including both healthy participants and patients representing 59 disease cohorts—to generate a unified pan-disease atlas serving as a high-resolution reference for human health and pathology.

By quantifying up to 5,416 circulating proteins using high-throughput affinity-based proteomics, the study provides an unprecedented dataset for examining how protein profiles vary across diverse disease states, demographic variables, and physiological conditions. Statistical modeling and machine learning analyses revealed both shared proteomic shifts common to multiple diseases and distinct molecular signatures specific to individual conditions. Such differentiation is critical for separating core inflammatory or systemic responses from protein alterations that may serve as reliable disease biomarkers.

The study additionally highlights that demographic variables such as age, sex, and body mass index (BMI) significantly influence circulating protein levels, underscoring the need to incorporate population stratification into biomarker discovery pipelines. Notably, the results demonstrate that each individual possesses a personalized proteomic signature that evolves over the lifespan: blood proteomes fluctuate more widely in childhood but stabilize in adulthood, offering a stable baseline against which pathological deviations may be detected more sensitively.

All data and analytical results have been made openly accessible through the Human Protein Atlas portal, enabling the broader scientific community to interrogate protein–disease relationships and accelerate precision medicine research. The publication underscores Prof. Dr. Gizem Dinler Doğanay’s integral role in advancing systems-level understanding of blood-based molecular signatures and contributes to strengthening Türkiye’s visibility in large-scale international biomedical research.

DOI: https://doi.org/10.1126/science.adx2678

5. Molecular Characterization of Antifungal Drug Resistance

In a recent international collaborative study with Assoc. Prof. Dr. Cecilia Geijer from Chalmers University of Technology (Sweden), published in the prestigious journal FEMS Yeast Research (Q2 in the Scopus index), Prof. Zeynep Petek Çakar’s group and collaborators developed and characterized an antimycin A-resistant Saccharomyces cerevisiae strain through evolutionary engineering as the Ph.D. study of Alican Topaloğlu. A portion of this work was conducted at Chalmers University of Technology by Ph.D. student Alican Topaloğlu, supported by an international FEMS (Federation of European Microbiological Societies) Research and Training Grant.

Prof. Çakar's research group at ITU Department of Molecular Biology & Genetics has been successfully improving resistance of yeast cells to various stress types. The employed evolutionary engineering approach induces significant physiological alterations in yeast. Therefore, identifying the metabolic and molecular differences between original and evolved strains is essential for developing microorganisms with improved industrial traits.

Antimycin A, an antifungal agent that inhibits mitochondrial respiration, provides a useful model for the growing clinical concern of antifungal drug resistance. Researchers have developed the first genetically stable, antimycin A-resistant S. cerevisiae strain using an evolutionary engineering strategy by applying gradually increasing antimycin A stress over long-term batch cultures. Whole-genome resequencing revealed two missense mutations in the PDR1 and PRP8 genes associated with the resistance phenotype. State-of-art gene modification technique CRISPR/Cas9 and subsequent cross-resistance tests confirmed that the PDR1 mutation alone is sufficient to confer antimycin A-resistance. Transcriptomic profiling further highlighted the central role of PDR1 gene in antifungal drug resistance and alterations in transmembrane transport, vesicular trafficking, and autophagy pathways.

DOI: https://doi.org/10.1093/femsyr/foaf062

6. Transforming Eggshell and Onion Membranes into High-Performance Enzyme Platforms: The First Comparative Study on Urease Immobilization

Researchers at Istanbul Technical University have conducted the first systematic comparative analysis demonstrating how waste-derived eggshell and onion membranes can be transformed into highly efficient, low-cost platforms for enzyme immobilization. Their findings are presented in the study titled “PEI-Treated Eggshell and Onion Membranes as Natural Supports for Urease Immobilization: A Performance and Stability Evaluation,” for which Assistant Professor Abdulhalim Kılıç of Istanbul Technical University serves as the corresponding author. The study offers a sustainable, technically rigorous approach to enhancing the stability of Jack Bean urease (JBU). This widely used industrial enzyme is often constrained by its limited stability in its free form.

By pretreating eggshell membrane (ESM) and onion membrane (OM) with polyethyleneimine (PEI), the team immobilized urease onto these naturally occurring biomaterials and assessed the resulting biocatalyst systems using morphological, kinetic, and stability-focused evaluations. Structural analyses showed that the porous, fibrous architecture of ESM supported higher enzyme loading, whereas OM—despite its more compact microstructure—provided superior operational robustness under thermal and storage stress.

Kinetic measurements revealed an expected increase in apparent Km due to diffusion restrictions introduced by immobilization, while both systems exhibited higher apparent Vmax as a result of greater local enzyme density on the membrane surfaces. Thermal and storage stability tests demonstrated that OM-immobilized urease maintained enzymatic activity up to 80°C and displayed notable dry-storage resilience. In contrast, ESM-supported urease excelled in reusability, preserving approximately 50% of its initial activity after four sequential cycles—double that of the OM system. Importantly, both biomaterial supports conferred substantial protection against Cu²⁺-induced inhibition, retaining more than 70% activity under conditions that completely inactivated free urease.

Overall, the study illustrates that eggshell and onion membranes are far more than biological waste—they function as versatile, sustainable, and cost-effective biomaterials capable of supporting high-performance enzyme immobilization. By merging environmental circularity with enhanced catalytic stability, this work establishes a promising foundation for developing durable biocatalyst platforms for industrial biotechnology, biosensing, and environmental applications.

DOI: https://doi.org/10.1021/acsomega.5c05782