GroEL ring separation and exchange in the chaperonin reaction

Yan, Xiao (2019): GroEL ring separation and exchange in the chaperonin reaction. Dissertation, LMU München: Fakultät für Chemie und Pharmazie

Abstract

The bacterial chaperonin GroEL and its cofactor GroES form a nano-cage for a single molecule of substrate protein (SP) to fold in isolation. GroEL and GroES undergo an ATP-regulated interaction cycle that governs the closing and opening of the folding cage. GroEL consists of two heptameric rings, stacked back-to-back, and displays intra-ring positive allosteric cooperativity and inter-ring negative allostery. Previous reports have suggested that ring separation and exchange can occur between the non-covalently bound rings of GroEL; however, the mechanism and physiological function of this phenomenon had yet to be explained. Here I show that GroEL undergoes transient ring separation, resulting in ring exchange between complexes. Through the ATPase cycling of GroEL/ES, ring separation is shown to occur upon ATP-binding to the trans-ring of the asymmetric GroEL:7ADP:GroES complex in the presence or absence of SP. Ring separation is a consequence of inter-ring negative allostery. To address the physiological function of this phenomenon, I created a novel mutant with the two rings connected by disulfide bonds. This GroEL mutant, unable to perform ring separation, is folding-active but populates symmetric GroEL:GroES2 complexes with GroES bound to both rings of GroEL, where both GroEL rings function simultaneously rather than sequentially. As a consequence, SP binding and release from the folding chamber is inefficient, and E. coli growth is impaired. My results suggest that transient ring separation is an integral part of the chaperonin mechanism to ensure sequential GroEL/ES cycling and effective SP folding.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: Protein folding, GroEL, GroES, Chaperonin, Fluorescence correlation spectroscopy, Dual-color fluorescence correlation spectroscopy, Molecular chaperone
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 540 Chemie
• Fakultäten: Fakultät für Chemie und Pharmazie
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 28. Januar 2019
• 1. Bericht­erstatter:in: Hartl, F. Ulrich