Long-read sequencing to detect full-length protein-protein interactions.
Publication/Presentation Date
7-17-2025
Abstract
Given the increased predictions on interactome size and demand for protein function information, methods for detecting protein-protein interactions remain a significant development area. The all-vs.-all sequencing (AVA-Seq) method utilizes a convergent fusion plasmid design to make two-hybrid technology amenable to next-generation sequencing. Here, we further innovate to take advantage of synthetic DNA technologies and Oxford Nanopore Technologies long-read sequencing improvements to allow us to determine full-length protein-protein interactions. We tested 3,115 human protein-protein pairs using this approach and recovered 159 protein-protein interactions from a set of 57 full-length human proteins. Fifteen of the 159 full-length protein-protein interactions matched known human interactions. When referencing a human gold standard set of interactions, eight full-length protein-protein interactions were recovered from an expected 28 interaction pairs (28.6%), a typical recovery rate for two-hybrid technologies. The AVA-Seq method, in combination with the ease of synthetic DNA production and the MinION platform, offers a low-cost, high-throughput alternative for determining protein-protein interactions, which can be utilized in research labs at all stages.
Volume
15
Issue
1
First Page
25967
Last Page
25967
ISSN
2045-2322
Published In/Presented At
Schaefer-Ramadan, S., Guan, Y., Ahmed, A. A., Aleksic, J., Elmagarmid, K. A., Syed, L. F., Mohamoud, Y. A., & Malek, J. A. (2025). Long-read sequencing to detect full-length protein-protein interactions. Scientific reports, 15(1), 25967. https://doi.org/10.1038/s41598-025-08549-3
Disciplines
Medicine and Health Sciences
PubMedID
40676061
Department(s)
Fellows and Residents
Document Type
Article