Structure and conformational cycle of a bacteriophage-encoded chaperonin

Abstract

Chaperonins are ubiquitous molecular chaperones found in all domains of life. They form ring-shaped complexes that assist in the folding of substrate proteins in an ATP-dependent reaction cycle. Key to the folding cycle is the transient encapsulation of substrate proteins by the chaperonin. Here we present a structural and functional characterization of the chaperonin gp146 ({phi}EL) from the phage EL of Pseudomonas aeruginosa. {phi}EL, an evolutionary distant homolog of bacterial GroEL, is active in ATP hydrolysis and prevents the aggregation of denatured protein in a nucleotide-dependent manner. However, {phi}EL failed to refold the encapsulation-dependent model substrate rhodanese and did not interact with E. coli GroES, the lid-shaped co-chaperone of GroEL. {phi}EL forms tetradecameric double-ring complexes, which dissociate into single rings in the presence of ATP. Crystal structures of {phi}EL (at 3.54 and 4.03 [A]) in presence of ATP*BeFx revealed two distinct single-ring conformational states, both with open access to the ring cavity. One state showed uniform ATP-bound subunit conformations (symmetric state), whereas the second combined distinct ATP- and ADP-bound subunit conformations (asymmetric state). Cryo-electron microscopy of apo-{phi}EL revealed a double-ring structure composed of rings in the asymmetric state (3.45 [A] resolution). We propose that the phage chaperonin undergoes nucleotide-dependent conformational switching between double- and single rings and functions in aggregation prevention without substrate protein encapsulation. Thus, {phi}EL may represent an evolutionary more ancient chaperonin prior to acquisition of the encapsulation mechanism.