Efficient ASK‐assisted system for expression and purification of plant F‐box proteins. (6th October 2017)

• Record Type: Journal Article
• Authors: Li, Haiou; Yao, Ruifeng; Ma, Sui; Hu, Shuai; Li, Suhua; Wang, Yupei; Yan, Chun; Xie, Daoxin; Yan, Jianbin
• Is Part Of: Plant journal. Volume 92:Number 4(2017)
• Page Start: 736
• Page End: 743
• Publication Date: 2017-10-06
• Subjects: COI1 -- SLY1 -- TIR1 -- ASK -- F‐box protein -- Technical Advance; Plant molecular biology -- Periodicals; Plant cells and tissues -- Periodicals; Botany -- Periodicals; 580
• DOI: 10.1111/tpj.13708
• Languages: English
• ISSNs: 0960-7412

Abstract

Summary: Ubiquitin‐mediated protein degradation plays an essential role in plant growth and development as well as responses to environmental and endogenous signals. F‐box protein is one of the key components of the SCF (SKP1‐CUL1‐F‐box protein) E3 ubiquitin ligase complex, which recruit specific substrate proteins for subsequent ubiquitination and 26S proteasome‐mediated degradation to regulate developmental processes and signaling networks. However, it is not easy to obtain purified F‐box proteins with high activity due to their unstable protein structures. Here, we found that Arabidopsis SKP‐like proteins (ASKs) can significantly improve soluble expression of F‐box proteins and maintain their bioactivity. We established an efficient ASK‐assisted method to express and purify plant F‐box proteins. The method meets a broad range of criteria required for the biochemical analysis or protein crystallization of plant F‐box proteins. Significance Statement: Plant F‐box proteins, as a key component of the SCF E3 ubiquitin ligase complex, play important roles in regulating plant growth and development during the entire plant life cycle. However, difficulties in obtaining purified F‐box proteins have affected their in‐depth study. In this study, we develop an effective ASK‐assisted baculovirus‐insect cell expression system that is suitable for the expression and purification of various F‐box proteins.