Minichaperone (GroEL191-345) mediated folding of MalZ proceeds by binding and release of native and functional intermediates.
TL;DR: Observations suggest that the minichaperone works by carrying out repeated cycles of binding aggregation-prone protein MalZ in a relatively compact conformation and in a partially folded but active state, and releasing them to attempt to fold in solution.
About: This article is published in Biochimica et Biophysica Acta.The article was published on 2018-09-01 and is currently open access. It has received 2 citations till now. The article focuses on the topics: GroEL & GroES.
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TL;DR: In this paper, the interaction of various htt ex1 constructs with the bacterial analog (GroEL) of the human chaperonin Hsp60 was investigated using fluorescence spectroscopy and electron and atomic force microscopy.
Abstract: Huntington's disease arises from polyQ expansion within the exon-1 region of huntingtin (htt ex1 ), resulting in an aggregation prone protein that accumulates in neuronal inclusion bodies. We investigate the interaction of various htt ex1 constructs with the bacterial analog (GroEL) of the human chaperonin Hsp60. Using fluorescence spectroscopy and electron and atomic force microscopy we show that GroEL inhibits fibril formation. The binding kinetics of htt ex1 constructs with intact GroEL and a mini-chaperone comprising the apical domain is characterized by relaxation-based NMR measurements. The lifetimes of the complexes range from 100-400 m s with equilibrium dissociation constants ( K D ) of ~1-2 mM. The binding interface is formed by the N-terminal amphiphilic region of htt ex1 (which adopts a partially helical conformation) and the H and I helices of the GroEL apical domain. Sequestration of monomeric htt ex1 by GroEL likely increases the critical concentration required for fibrillization.
7 citations
20 Jan 2020
TL;DR: The main events in chaperone-assisted protein folding are the binding and ligand-induced release of substrate proteins and the time of GroES-induced dissociation of a denatured protein from the GroEL surface was found to be much shorter than the proposed time of the GroES ATPase cycle.
Abstract: The main events in chaperone-assisted protein folding are the binding and ligand-induced release of substrate proteins. Here, we studied the location of denatured proteins previously bound to the GroEL chaperonin resulting from the action of the GroES co-chaperonin in the presence of Mg-ATP. Fluorescein-labeled denatured proteins (α-lactalbumin, lysozyme, serum albumin, and pepsin in the presence of thiol reagents at neutral pH, as well as an early refolding intermediate of malate dehydrogenase) were used to reveal the effect of GroES on their interaction with GroEL. Native electrophoresis has demonstrated that these proteins tend to be released from the GroEL-GroES complex. With the use of biotin- and fluorescein-labeled denatured proteins and streptavidin fused with luciferase aequorin (the so-called streptavidin trap), the presence of denatured proteins in bulk solution after GroES and Mg-ATP addition has been confirmed. The time of GroES-induced dissociation of a denatured protein from the GroEL surface was estimated using the stopped-flow technique and found to be much shorter than the proposed time of the GroEL ATPase cycle.
4 citations
References
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TL;DR: The crystal structure of Escherichia coli GroEL shows a porous cylinder of 14 subunits made of two nearly 7-fold rotationally symmetrical rings stacked back-to-back with dyad symmetry.
Abstract: The crystal structure of Escherichia coli GroEL shows a porous cylinder of 14 subunits made of two nearly 7-fold rotationally symmetrical rings stacked back-to-back with dyad symmetry. The subunits consist of three domains: a large equatorial domain that forms the foundation of the assembly at its waist and holds the rings together; a large loosely structured apical domain that forms the ends of the cylinder; and a small slender intermediate domain that connects the two, creating side windows. The three-dimensional structure places most of the mutationally defined functional sites on the channel walls and its outward invaginations, and at the ends of the cylinder.
1,285 citations
TL;DR: This review focuses on recent advances in understanding the mechanisms of chaperone action in promoting and regulating protein folding and on the pathological consequences of protein misfolding and aggregation.
Abstract: The biological functions of proteins are governed by their three-dimensional fold. Protein folding, maintenance of proteome integrity, and protein homeostasis (proteostasis) critically depend on a complex network of molecular chaperones. Disruption of proteostasis is implicated in aging and the pathogenesis of numerous degenerative diseases. In the cytosol, different classes of molecular chaperones cooperate in evolutionarily conserved folding pathways. Nascent polypeptides interact cotranslationally with a first set of chaperones, including trigger factor and the Hsp70 system, which prevent premature (mis)folding. Folding occurs upon controlled release of newly synthesized proteins from these factors or after transfer to downstream chaperones such as the chaperonins. Chaperonins are large, cylindrical complexes that provide a central compartment for a single protein chain to fold unimpaired by aggregation. This review focuses on recent advances in understanding the mechanisms of chaperone action in promoting and regulating protein folding and on the pathological consequences of protein misfolding and aggregation.
1,249 citations
"Minichaperone (GroEL191-345) mediat..." refers background in this paper
...Cells circumvent this problem in part by the expression of molecular chaperones, which act by binding to nascent polypeptides or by providing protected environments where proteins can fold without interacting with other aggregation-prone intermediates (14-17)...
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TL;DR: Genetic and biochemical analysis shows that several distinct chaperone systems, including Hsp70 and the cylindrical chaperonins, assist the folding of proteins upon translation in the cytosol of both prokaryotic and eukaryotic cells.
Abstract: ▪ Abstract Recent years have witnessed dramatic advances in our understanding of how newly translated proteins fold in the cell and the contribution of molecular chaperones to this process. Folding in the cell must be achieved in a highly crowded macromolecular environment, in which release of nonnative polypeptides into the cytosolic solution might lead to formation of potentially toxic aggregates. Here I review the cellular mechanisms that ensure efficient folding of newly translated proteins in vivo. De novo protein folding appears to occur in a protected environment created by a highly processive chaperone machinery that is directly coupled to translation. Genetic and biochemical analysis shows that several distinct chaperone systems, including Hsp70 and the cylindrical chaperonins, assist the folding of proteins upon translation in the cytosol of both prokaryotic and eukaryotic cells. The cellular chaperone machinery is specifically recruited to bind to ribosomes and protects nascent chains and foldi...
1,149 citations
TL;DR: A new view of protein folding is emerging, whereby the energy landscapes that proteins navigate during folding in vivo may differ substantially from those observed during refolding in vitro.
Abstract: Most proteins must fold into unique three-dimensional structures to perform their biological functions. In the crowded cellular environment, newly synthesized proteins are at risk of misfolding and forming toxic aggregate species. To ensure efficient folding, different classes of molecular chaperones receive the nascent protein chain emerging from the ribosome and guide it along a productive folding pathway. Because proteins are structurally dynamic, constant surveillance of the proteome by an integrated network of chaperones and protein degradation machineries is required to maintain protein homeostasis (proteostasis). The capacity of this proteostasis network declines during aging, facilitating neurodegeneration and other chronic diseases associated with protein aggregation. Understanding the proteostasis network holds the promise of identifying targets for pharmacological intervention in these pathologies.
1,009 citations
"Minichaperone (GroEL191-345) mediat..." refers background in this paper
...Cells circumvent this problem in part by the expression of molecular chaperones, which act by binding to nascent polypeptides or by providing protected environments where proteins can fold without interacting with other aggregation-prone intermediates (14-17)...
[...]
TL;DR: Using recent studies, can the authors begin to search for trends which may lead to a better understanding of the protein folding process, and stable intermediates are not a prerequisite for the fast, efficient folding of proteins.
895 citations