An essential nonredundant role for mycobacterial DnaK in native protein folding.

Allison Fay,Michael S Glickman,Patrick H Viollier

doi:10.1371/journal.pgen.1004516

Allison Fay, Michael S Glickman + Show 1 more

Open Access

https://doi.org/10.1371/journal.pgen.1004516

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Journal: PLoS Genetics	Publication Date: Jul 24, 2014
Citations: 123	License type: CC BY 4.0

Affiliation: Memorial Sloan Kettering Cancer Center

Abstract

Protein chaperones are essential in all domains of life to prevent and resolve protein misfolding during translation and proteotoxic stress. HSP70 family chaperones, including E. coli DnaK, function in stress induced protein refolding and degradation, but are dispensable for cellular viability due to redundant chaperone systems that prevent global nascent peptide insolubility. However, the function of HSP70 chaperones in mycobacteria, a genus that includes multiple human pathogens, has not been examined. We find that mycobacterial DnaK is essential for cell growth and required for native protein folding in Mycobacterium smegmatis. Loss of DnaK is accompanied by proteotoxic collapse characterized by the accumulation of insoluble newly synthesized proteins. DnaK is required for solubility of large multimodular lipid synthases, including the essential lipid synthase FASI, and DnaK loss is accompanied by disruption of membrane structure and increased cell permeability. Trigger Factor is nonessential and has a minor role in native protein folding that is only evident in the absence of DnaK. In unstressed cells, DnaK localizes to multiple, dynamic foci, but relocalizes to focal protein aggregates during stationary phase or upon expression of aggregating peptides. Mycobacterial cells restart cell growth after proteotoxic stress by isolating persistent DnaK containing protein aggregates away from daughter cells. These results reveal unanticipated essential nonredunant roles for mycobacterial DnaK in mycobacteria and indicate that DnaK defines a unique susceptibility point in the mycobacterial proteostasis network.

Highlights

Proper protein folding is essential for all organisms and assures that the primary sequence of the polypeptide forms its functional tertiary and quaternary structures
In E. coli, DnaK is nonessential for native protein folding because of redundancy with Trigger Factor, which associates with proteins soon after emergence from the ribosome [11]
DnaK is essential for growth of M. smegmatis To study the function of mycobacterial DnaK, we attempted to delete dnaK from the M. smegmatis chromosome

Summary

Introduction

Proper protein folding is essential for all organisms and assures that the primary sequence of the polypeptide forms its functional tertiary and quaternary structures. Chaperones are required to assure proper protein folding and prevent protein aggregation, which can occur as hydrophobic segments of the protein emerge from the ribosome. DnaK is a central hub for protein folding, shuttling misfolded peptides to other chaperones and proteases for resolution, a function that is essential during the protein denaturation that occurs during heat shock [6,7,8,9]. In E. coli, DnaK is nonessential for native protein folding because of redundancy with Trigger Factor, which associates with proteins soon after emergence from the ribosome [11]. Examination of proteins that interact with DnaK indicates that most client proteins that require DnaK for proper folding and/or stability are largely non-essential, suggesting that loss of function of these proteins in the absence of DnaK does not impact viability [7]. In the absence of both DnaK and TF, the E. coli cell suffers proteostasis collapse characterized by global insolubility of nascent proteins [7]

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