Assembly and Disassembly of GroEL and GroES Complexes

Abstract

The assembly of chaperonin subunits into heptameric ring structures is a hallmark common to this group of proteins. In GroEL two of these rings are symmetrically stacked, resulting in a tetradecameric double-ring cylinder which is the functional form of the chaperonin. In this structure, unfolded substrate proteins bind to the apical subunit domains at the inner surface of the cylinder, whereas adenosine 5’-triphosphate (ATP) binding and hydrolysis occur at the level of the central subunit domains (1). GroES subunits assemble into a single heptameric ring, which can bind asymmetrically on top of a GroEL ring (2). Interestingly, the intracellular assembly of GroEL-type chaperonins appears to be a chaperone-dependent process itself and requires functional preformed chaperonin complexes (3). The intracellular folding pathway of GroES has not been analyzed, but in vitro the complex is able to assemble spontaneously from monomeric subunits (4,5). Cell-free assembly of GroEL in a Mg-ATP-dependent manner, starting from monomeric subunits, has been established, too (6,7). Like with GroES, in vitro assembly of GroEL is a reversible process, and depending on the conditions, dissociation of complexes can occur. The formation of these oligomeric structures poses thus an interesting folding problem per se. However, in addition to providing an interesting model system for folding and assembly studies, controlled (dis)assembly of chaperonins has also been found to be helpful in generating highly purified chaperonin complexes (8). A common problem during purification of GroEL is its contamination with bound substrate proteins, which are difficult to remove by conventional purification steps (see Chapter 3). Disassembly of GroEL into folded monomers and subsequent reassembly can result in functional chaperonin complexes, which are largely devoid of bound substrates. In a different application, site-directed mutagenesis studies of GroEL, the readiness with which the complex dissociates and reassembles, has been assayed to obtain information about the role of certain amino acid residues in the stability of the complex and in its oligomerization (9,10).