Aliarcobacter butzleri is an emerging foodborne and zoonotic pathogen, yet many of its encoded proteins remain functionally uncharacterized. This lack of annotation limits understanding of its molecular mechanisms and hampers the identification of novel therapeutic targets. In this study, we systematically performed functional annotation of essential hypothetical proteins from the BNI-3166 strain using an integrative-in-silico approach to uncover potential drug and vaccine candidates. 2,367 protein-coding sequences were retrieved from the RefSeq database and were identified 356 as hypothetical proteins. Using BLASTp, we screened these HPs against the Database of Essential Genes and the human proteome to identify essential non-homologous proteins, resulting in 20 ENH candidates. Functional annotation was performed using several domain-based databases, including Pfam, InterPro, SMART, and SUPERFAMILY. Subsequently, physicochemical properties were analyzed and predicted subcellular localization using PSORTb and CELLO. To assess druggability, the ChEMBL database was used. Virulence factors using VFDB, VICMpred, and VirulentPred 2.0 were also predicted. Gene Ontology annotations were generated via ARGOT2.5. Furthermore, we explored protein-protein interactions using STRING and predicted tertiary structures with AlphaFold3. Moreover, Ligand binding pockets were predicted using PrankWeb, and antigenicity of vaccine candidates was assessed using VaxiJen v2.0. We identified 20 essential non-homologous hypothetical proteins, of which 10 were confidently annotated based on conserved domain analysis. These proteins were classified as enzymes, binding proteins, transporters, regulatory proteins, and potential virulence factors. Among them, eight exhibited characteristics of promising drug targets, while two showed potential as vaccine candidates based on subcellular localization. Druggability analysis revealed that nine proteins had no similarity to known drug targets, suggesting novel therapeutic potential. Predicted 3D structures generated using AlphaFold3 yielded pTM scores ranging from 0.44 to 0.92, indicating acceptable to high modeling confidence. Ligand binding site analysis confirmed druggability in six candidates, and antigenicity screening identified one protein as a potential vaccine target. This study provides a computational framework for identifying functionally important proteins in A. butzleri BNI-3166 and highlights novel therapeutic candidates for experimental validation, offering new directions in drug and vaccine development against this underexplored pathogen.
Key words: Aliarcobacter butzleri, Drug Target Identification, Functional Annotation, Hypothetical Proteins, In Silico Analysis
Received: 08.07.2025; Accepted: 01.09.2025; Early view: 24.09.2025 Published: 10.01.2026
DOI: 10.62063/ecb-66
Citation: Paul, S., Barua, S., & Barua, J.D. (2026). In-silico functional annotation and structural characterization of hypothetical proteins from Aliarcobacter butzleri BNI-3166: Insights into novel virulence and drug targets. The European chemistry and biotechnology journal, 5, 22-39. https://doi.org/10.62063/ecb-66
The copyrights of the studies published in The European Chemistry and Biotechnology Journal (EUCHEMBIOJ) belong to their authors
This article is distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0)(https://creativecommons.org/licenses/by-nc/4.0/).
If you are running Windows 8, 8.1, or even newer operating systems and want to avoid the legal and security hazards of cracked software, several highly capable, free, and safe alternatives exist.
Preactivated software, often distributed via torrent sites like 4realtorrentz, implies the software has been pre-patched or cracked. Users do not need to enter a license key or use a keygen. Why Users Choose This Route: Immediate Access: No payment or registration required.
The search query "stardock start8 156 preactivated 4realtorrentz crack better" is composed of several distinct elements used by piracy networks:
A lightweight, free utility that brings back classic Windows taskbars and Start menus from older versions of Windows. Conclusion
Searching for "Stardock Start8 156 preactivated 4realtorrentz crack" might seem like a quick way to get a premium tool for free, but the risks heavily outweigh the rewards. The high probability of infecting your system with malware, losing your personal data, or causing operating system crashes makes cracked software a dangerous choice.
If you are running Windows 8, 8.1, or even newer operating systems and want to avoid the legal and security hazards of cracked software, several highly capable, free, and safe alternatives exist.
Preactivated software, often distributed via torrent sites like 4realtorrentz, implies the software has been pre-patched or cracked. Users do not need to enter a license key or use a keygen. Why Users Choose This Route: Immediate Access: No payment or registration required.
The search query "stardock start8 156 preactivated 4realtorrentz crack better" is composed of several distinct elements used by piracy networks:
A lightweight, free utility that brings back classic Windows taskbars and Start menus from older versions of Windows. Conclusion
Searching for "Stardock Start8 156 preactivated 4realtorrentz crack" might seem like a quick way to get a premium tool for free, but the risks heavily outweigh the rewards. The high probability of infecting your system with malware, losing your personal data, or causing operating system crashes makes cracked software a dangerous choice.