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/).
Arun Vijay's action sequences are a major draw, making it a "must-watch" for thriller fans.
Kuttram 23 Tamilyogi: Navigating the Legality, Impact, and Legacy of the Medical Thriller
Tamilyogi is a notorious, piracy-based website that illegally distributes copyrighted content. It specializes in hosting a vast collection of movies, primarily in Tamil, but also in other South Indian languages like Telugu and Malayalam, as well as dubbed versions of Hindi and Hollywood films. It operates by making these movies available for free streaming and download, often in high-definition formats like 720p or 1080p, very soon after their official release.
Kuttram 23 stands as a testament to what Kollywood can achieve when sharp writing meets dedicated performances. Engaging with this cinematic piece through queries like "Kuttram 23 Tamilyogi" undermines the very industry that crafts these stories. By opting for official OTT channels and legal theater releases, viewers ensure that creators are fairly compensated, paving the way for more high-quality medical thrillers in the future.
Searchers are usually looking for the full 2-hour-plus runtime to experience the complete investigation.
The film kicks off at a frantic pace when a distressed pregnant woman enters a church in Chennai, only for the priest inside to be shot dead in his confession booth. The woman and her car vanish shortly after. (Arun Vijay), a sharp and fiercely dedicated police officer, is assigned to crack the high-profile missing person case. Kuttram 23 (2017) - Movie | Reviews, Cast & Release Date
Accessing movies through Tamilyogi is a violation of copyright laws. The platform does not hold the necessary licenses or permissions from the original creators, making its entire operation illegal. In India, such piracy is combated under the Copyright Act, the Information Technology Act, and the Cinematograph Act.
Piracy constitutes copyright infringement, which is illegal in India and many other countries. Under Sections 63 and 63A of the Indian Copyright Act, anyone knowingly infringing copyright can face:
Using piracy sites like TamilYogi to watch content is generally discouraged for several reasons: Device Safety: Security experts from
Arun Vijay's action sequences are a major draw, making it a "must-watch" for thriller fans.
Kuttram 23 Tamilyogi: Navigating the Legality, Impact, and Legacy of the Medical Thriller
Tamilyogi is a notorious, piracy-based website that illegally distributes copyrighted content. It specializes in hosting a vast collection of movies, primarily in Tamil, but also in other South Indian languages like Telugu and Malayalam, as well as dubbed versions of Hindi and Hollywood films. It operates by making these movies available for free streaming and download, often in high-definition formats like 720p or 1080p, very soon after their official release.
Kuttram 23 stands as a testament to what Kollywood can achieve when sharp writing meets dedicated performances. Engaging with this cinematic piece through queries like "Kuttram 23 Tamilyogi" undermines the very industry that crafts these stories. By opting for official OTT channels and legal theater releases, viewers ensure that creators are fairly compensated, paving the way for more high-quality medical thrillers in the future.
Searchers are usually looking for the full 2-hour-plus runtime to experience the complete investigation.
The film kicks off at a frantic pace when a distressed pregnant woman enters a church in Chennai, only for the priest inside to be shot dead in his confession booth. The woman and her car vanish shortly after. (Arun Vijay), a sharp and fiercely dedicated police officer, is assigned to crack the high-profile missing person case. Kuttram 23 (2017) - Movie | Reviews, Cast & Release Date
Accessing movies through Tamilyogi is a violation of copyright laws. The platform does not hold the necessary licenses or permissions from the original creators, making its entire operation illegal. In India, such piracy is combated under the Copyright Act, the Information Technology Act, and the Cinematograph Act.
Piracy constitutes copyright infringement, which is illegal in India and many other countries. Under Sections 63 and 63A of the Indian Copyright Act, anyone knowingly infringing copyright can face:
Using piracy sites like TamilYogi to watch content is generally discouraged for several reasons: Device Safety: Security experts from