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Efficient Catalysis of Protein Folding by GroEL/ES of the Obligate Chaperonin Substrate MetF.
J Mol Biol. 2020 03 27; 432(7):2304-2318.JM

Abstract

The cylindrical chaperonin GroEL and its cofactor GroES mediate ATP-dependent protein folding in Escherichia coli by transiently encapsulating non-native substrate in a nano-cage formed by the GroEL ring cavity and the lid-shaped GroES. Mechanistic studies of GroEL/ES with heterologous protein substrates suggested that the chaperonin is inefficient, typically requiring multiple ATP-dependent encapsulation cycles with only a few percent of protein folded per cycle. Here we analyzed the spontaneous and chaperonin-assisted folding of the essential enzyme 5,10-methylenetetrahydrofolate reductase (MetF) of E. coli, an obligate GroEL/ES substrate. We found that MetF, a homotetramer of 33-kDa subunits with (β/α)8 TIM-barrel fold, populates a kinetically trapped folding intermediate(s) (MetF-I) upon dilution from denaturant that fails to convert to the native state, even in the absence of aggregation. GroEL/ES recognizes MetF-I and catalyzes rapid folding, with ~50% of protein folded in a single round of encapsulation. Analysis by hydrogen/deuterium exchange at peptide resolution showed that the MetF subunit folds to completion in the GroEL/ES nano-cage and binds its cofactor flavin adenine dinucleotide. Rapid folding required the net negative charge character of the wall of the chaperonin cavity. These findings reveal a remarkable capacity of GroEL/ES to catalyze folding of an endogenous substrate protein that would have coevolved with the chaperonin system.

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Publisher Full Text (DOI)

Authors+Show Affiliations

Singh AK
Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82159 Martinsried, Germany.
Balchin D
Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82159 Martinsried, Germany.
Imamoglu R
Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82159 Martinsried, Germany.
Hayer-Hartl M
Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82159 Martinsried, Germany. Electronic address: mhartl@biochem.mpg.de.
Hartl FU
Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82159 Martinsried, Germany. Electronic address: uhartl@biochem.mpg.de.

MeSH

5,10-Methylenetetrahydrofolate Reductase (FADH2)Adenosine TriphosphateCatalysisEscherichia coliEscherichia coli ProteinsHeat-Shock ProteinsKineticsModels, MolecularProtein ConformationProtein FoldingThermodynamics

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

32135190

Citation

Singh, Amit K., et al. "Efficient Catalysis of Protein Folding By GroEL/ES of the Obligate Chaperonin Substrate MetF." Journal of Molecular Biology, vol. 432, no. 7, 2020, pp. 2304-2318.
Singh AK, Balchin D, Imamoglu R, et al. Efficient Catalysis of Protein Folding by GroEL/ES of the Obligate Chaperonin Substrate MetF. J Mol Biol. 2020;432(7):2304-2318.
Singh, A. K., Balchin, D., Imamoglu, R., Hayer-Hartl, M., & Hartl, F. U. (2020). Efficient Catalysis of Protein Folding by GroEL/ES of the Obligate Chaperonin Substrate MetF. Journal of Molecular Biology, 432(7), 2304-2318. https://doi.org/10.1016/j.jmb.2020.02.031
Singh AK, et al. Efficient Catalysis of Protein Folding By GroEL/ES of the Obligate Chaperonin Substrate MetF. J Mol Biol. 2020 03 27;432(7):2304-2318. PubMed PMID: 32135190.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Efficient Catalysis of Protein Folding by GroEL/ES of the Obligate Chaperonin Substrate MetF. AU - Singh,Amit K, AU - Balchin,David, AU - Imamoglu,Rahmi, AU - Hayer-Hartl,Manajit, AU - Hartl,F Ulrich, Y1 - 2020/03/02/ PY - 2020/01/18/received PY - 2020/02/25/revised PY - 2020/02/25/accepted PY - 2020/3/7/pubmed PY - 2020/8/28/medline PY - 2020/3/6/entrez KW - Chaperonins KW - GroEL KW - Hydrogen/deuterium exchange KW - MetF KW - Single molecule spectroscopy SP - 2304 EP - 2318 JF - Journal of molecular biology JO - J Mol Biol VL - 432 IS - 7 N2 - The cylindrical chaperonin GroEL and its cofactor GroES mediate ATP-dependent protein folding in Escherichia coli by transiently encapsulating non-native substrate in a nano-cage formed by the GroEL ring cavity and the lid-shaped GroES. Mechanistic studies of GroEL/ES with heterologous protein substrates suggested that the chaperonin is inefficient, typically requiring multiple ATP-dependent encapsulation cycles with only a few percent of protein folded per cycle. Here we analyzed the spontaneous and chaperonin-assisted folding of the essential enzyme 5,10-methylenetetrahydrofolate reductase (MetF) of E. coli, an obligate GroEL/ES substrate. We found that MetF, a homotetramer of 33-kDa subunits with (β/α)8 TIM-barrel fold, populates a kinetically trapped folding intermediate(s) (MetF-I) upon dilution from denaturant that fails to convert to the native state, even in the absence of aggregation. GroEL/ES recognizes MetF-I and catalyzes rapid folding, with ~50% of protein folded in a single round of encapsulation. Analysis by hydrogen/deuterium exchange at peptide resolution showed that the MetF subunit folds to completion in the GroEL/ES nano-cage and binds its cofactor flavin adenine dinucleotide. Rapid folding required the net negative charge character of the wall of the chaperonin cavity. These findings reveal a remarkable capacity of GroEL/ES to catalyze folding of an endogenous substrate protein that would have coevolved with the chaperonin system. SN - 1089-8638 UR - https://www.unboundmedicine.com/medline/citation/32135190/Efficient_Catalysis_of_Protein_Folding_by_GroEL/ES_of_the_Obligate_Chaperonin_Substrate_MetF_ DB - PRIME DP - Unbound Medicine ER -