IN SILICO STRUCTURAL AND SEQUENCE ANALYSIS OF DUF724 PROTEIN DOMAIN

Proteins containing the Domain of Unknown Function 724 (DUF724) are found exclusively in plants, conserved across monocots and dicots. Typically, multiple DUF724 members occur per organism: five in rice (O. sativa), ten in Arabidopsis thaliana (AtDuf1-10), and fourteen in wheat (T. aestivum). Most DUF724 members feature Agenet domains, histone readers. Thus, DUF724 is hypothesized to be involved in plant epigenetic control. Furthermore, epigenome editing is a highly promising area in crop improvement, as it facilitates the transmission of epigenetic marks. This research project aims to computationally study the structure and sequence of DUF724 across plant homologs to help discover conserved traits and provide insights into possible DUF724-DUF724 interactions. Structural predictions were performed on ten AtDuf’s (Uniprot entries DUF1-ARATH to DUF10-ARATH) using AlphaFold3 (AF3) with standard parameters, complemented by Python analyses (AlphaJudge library pipeline for model metrics; K-means clustering of mRNA pattern expression data via Scikit-learn and Pandas) for biological consistencies. AF3 predictions of DUF724 show an N-terminal helix-turn-helix-like motif (HTH), which could participate in homodimerization, and a C-terminal antiparallel dimeric coiled coil (CC) region, possibly facilitating filament formation, supported by in vitro studies from our group or Cao et al. Sequence analysis showed AtDuf’s can be classified into groups based on sequence similarity. A conserved residue position seems determinant for HTH homodimer conformation, for reasons being investigated. AF3 confidence metrics and stereochemical parameters suggested a possible heterodimerization between AtDuf’s, supported by tissue expression data from Cao et al. and our clustering analysis. Finally, phylogenetic tree, conservation, and ancient sequence reconstruction analyses are underway on 2,000+ InterPro DUF724 entries (IPR007930) to understand gene duplication events. A better characterization of DUF724 structure and interactions will expand knowledge of its potential functions and may enable agricultural innovations.