Method for preparing multisubunit SCF E3 ligase with fusion protein through in vitro reconstitution, and use of multisubunit SCF E3 ligase

US12359186B2 · kind B2 · utility

Assignee

• Institute of Genetics and Developmental Biology, CAS · CN

Inventors

• Jiayang Li · Tangxia, CN
• Bing Wang · San Jose, US
• Huihui Liu · Beijing, CN
• Hong Yu · Seojong-myeon, KR
• Xiangbing Meng · Beijing, CN
• Guifu Liu · Beijing, CN
• Mingjiang Chen · Beijing, CN
• Yanhui Jing · Beijing, CN

Key dates

• Filing date: May 25, 2023
• Grant date: Jul 15, 2025
• Expiry date: Jul 13, 2043

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C07K14/4702
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

The disclosure discloses a method for preparing a multisubunit SCF E3 ligase with a fusion protein through in vitro reconstitution, and a use of the multisubunit SCF E3 ligase. The method for preparing a multisubunit SCF E3 ligase with a fusion protein through in vitro reconstitution disclosed by the disclosure includes: subjecting an engineered SKP1-Cullin1-RBX1 fusion protein (referred to as eSCR) to a reaction with an F-box protein in a reaction system to obtain the multisubunit SCF E3 ligase, where the eSCR fusion protein has an amino acid sequence from position 10 to position 1062 of SEQ ID NO: 2. Experimental results show that different multisubunit SCF E3 ligases are successfully prepared with the eSCR fusion protein through in vitro reconstitution in the disclosure; the reconstituted multisubunit E3 ligase has biological activity.