A rapid and efficient extraction method for plant micropeptides

Le résumé fourni par la source

Micropeptides are proteins with molecular weights below 10 kDa that regulate many biological processes. However, their extraction efficiency remains insufficient for many plant species, particularly those with complex chemical compositions. Here, we present a rapid and efficient method for plant micropeptide extraction. Four extraction buffers designated M1 to M4 were developed. Comparative analysis showed that 1% trifluoroacetic acid provided the most suitable solvent for micropeptide re-dissolution. Based on these findings, we established an optimized method that integrated extraction with a buffer supplemented with hydrochloric acid, phenol extraction, acetone precipitation, re-dissolution in 1% trifluoroacetic acid, detection via tricine-SDS-PAGE, molecular weight cutoff filtration, and identification by LC-MS/MS. Tricine-SDS-PAGE and Orbitrap LC-MS/MS analysis showed that buffer M3 provided the highest micropeptide abundance and identification efficiency. Buffer M3 contained 800 mM sucrose, 15 mM Triton X-100, 250 mM β-mercaptoethanol, 10 mM Tris-HCl (pH 8.0), 100 mM EDTA, 50 mM borax, and 50 mM vitamin C. The optimization of hydrochloric acid concentration further improved the extraction efficiency in different plant species. The M3-based extraction method performed well in other natural rubber-producing plants ( Eucommia ulmoides and Hevea brasiliensis ), model plants ( Arabidopsis thaliana , Oryza sativa , and Nicotiana benthamiana ), the halophyte plant Sesuvium portulacastrum , the polyphenol-rich plant Camellia sinensis , and the polysaccharide-rich plant Saccharum officinarum . The method contains buffer M3 with optimized hydrochloric acid concentrations and 1% trifluoroacetic acid re-dissolution, offering a rapid, efficient, and broadly applicable strategy for micropeptide isolation and supports large-scale micropeptidomic studies across diverse plant species.

Ce résumé expose les affirmations des auteurs. BNTIC ne l’interprète pas comme une validation indépendante des résultats.