In E. coli cells, rescue of non-native proteins and promotion of native state structure is assisted by the chaperonin GroEL. An important key to this activity lies in the structure of the apical domain of GroEL (GroEL-AD) (residue 191-376), which recognizes and binds non-native protein molecules through hydrophobic interactions. In this study, we investigated the effects of GroEL-AD on the aggr...

Although a cis mechanism of GroEL-mediated protein folding, occurring inside a hydrophilic chamber encapsulated by the co-chaperonin GroES, has been well documented, recently the GroEL-GroES-mediated folding of aconitase, a large protein (82 kDa) that could not be encapsulated, was described. This process required GroES binding to the ring opposite the polypeptide (trans) to drive release and p...

Upon dilution, D-glyceraldehyde-3-phosphate dehydrogenase (GADPH) that has been fully inactivated, but only partially unfolded, in dilute guanidine hydrochloride (GuHCl) recovers activity completely. The fully unfolded enzyme, however, is re-activated only to a limited extent after dilution, and refolds rapidly in a burst phase to a partially folded intermediate characterized by increases in bo...

Chaperonin-assisted protein folding proceeds through cycles of ATP binding and hydrolysis by the large chaperonin GroEL, which undergoes major allosteric rearrangements. Interaction between the two back-to-back seven-membered rings of GroEL plays an important role in regulating binding and release of folding substrates and of the small chaperonin GroES. Using cryo-electron microscopy, we have o...

A 2.7 kb DNA fragment encoding the 60 kDa common antigen (CA) and a 13 kDa protein of Legionella micdadei was sequenced. Two open reading frames of 57,677 and 10,456 Da were identified, corresponding to the heat shock proteins GroEL and GroES, respectively. Typical -35, -10, and Shine-Dalgarno heat shock expression signals were identified upstream of the L. micdadei groEL gene. Further upstream...

Mirror-image proteins (composed of D-amino acids) are promising therapeutic agents and drug discovery tools, but as synthesis of larger D-proteins becomes feasible, a major anticipated challenge is the folding of these proteins into their active conformations. In vivo, many large and/or complex proteins require chaperones like GroEL/ES to prevent misfolding and produce functional protein. The a...

We have studied the ATP-induced allosteric structural transition of GroEL using small angle X-ray scattering and fluorescence spectroscopy in combination with a stopped-flow technique. With X-ray scattering one can clearly distinguish the three allosteric states of GroEL, and the kinetics of the transition of GroEL induced by 85 microM ATP have been observed directly by stopped-flow X-ray scatt...

Molecular chaperones fold many proteins and their mutated versions in a cell and can sometimes buffer the phenotypic effect of mutations that affect protein folding. Unanswered questions about this buffering include the nature of its mechanism, its influence on the genetic variation of a population, the fitness trade-offs constraining this mechanism, and its role in expediting evolution. Answer...

Several approaches were used to study the role of GroEL, the prototype chaperonin, in the nitrogen fixation (nif) system. An Escherichia coli groEL mutant transformed with the Klebsiella pneumoniae nif gene cluster accumulated very low to nondetectable levels of nitrogenase components compared with the isogenic wild-type strain or the mutant cotransformed with the wild-type groE operon. In K. p...

Bacterial chaperonin, GroEL, together with its co-chaperonin, GroES, facilitates the folding of a variety of polypeptides. Experiments suggest that GroEL stimulates protein folding by multiple cycles of binding and release. Misfolded proteins first bind to an exposed hydrophobic surface on GroEL. GroES then encapsulates the substrate and triggers its release into the central cavity of the GroEL...