DnaK Chaperone Takes Part in Folding but Not in Refolding of Thermal Inactivated Proteins in Bacillus subtilis
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DnaK Chaperone Takes Part in Folding but Not in Refolding of Thermal Inactivated Proteins in Bacillus subtilis E. Yu. Gnuchikha, c, *, I. V. Manukhovd, and G. B. Zavilgelskyb, c, ** a State Research Institute of Genetics and Selection of Industrial Microorganisms of the National Research Centre “Kurchatov Institute”, Kurchatov Genomic Center, Moscow, 117545 Russia b State Research Institute of Genetics and Selection of Industrial Microorganisms of the National Research Centre “Kurchatov Institute”, Moscow, 117545 Russia c National Research Centre “Kurchatov Institute”, Moscow, 123182 Russia d Laboratory of Molecular Genetics, Moscow Institute of Physics and Technology, Dolgoprudny, Moscow Region, 141700 Russia *e-mail: [email protected] **e-mail: [email protected]
Received November 7, 2019; revised December 12, 2019; accepted December 18, 2019
Abstract—The role of the DnaKJE and Trigger Factor (TF) chaperones in folding and refolding of proteins in Bacillus subtilis is studied. Bacterial luciferases of Photobacterium leiognathi and Photorhabdus luminescens as protein substrates have been used. It is shown that DnaKJE takes part in folding but not in refolding of the thermal inactivated proteins. It is shown that TF takes part in the synthesis of thermolabile P. leiognathi luciferase, but significantly decreases the level of synthesis of thermostable P. luminescens luciferase. Keywords: folding, refolding, chaperone, DnaKJE, Trigger Factor, streptomycin, mistranslation, bacterial luciferase DOI: 10.1134/S1022795420090070
INTRODUCTION In gram-negative bacteria Escherichia coli, chaperone DnaK-DnaJ-GrpE (DnaKJE) plays a key role in the process of monitoring product quality (proteins), destroying protein aggregates, and restoring native structures of misfolded and denatured (unfolded) proteins (refolding) [1–7]. In addition, DnaKJE participates together with the chaperone Trigger Factor (TF) in the formation of the native structure of the protein macromolecule during synthesis on the ribosome (folding) [8–12]. In gram-positive bacteria, the regulation of expression of chaperone proteins has been studied [13–15], as well as the effect of chaperones on the survival of bacteria at elevated (stress) temperatures [16]. It is shown that chaperon DnaK of Bacillus subtilis complements the ability to grow at high temperature (40°C) the mutant strain E. coli ΔdnaK [17]. It is shown that for B. subtilis the mechanism of regulation of heat shock gene expression differs significantly from that in cells of E. coli. Regarding the participation of chaperones in the processes of folding and refolding of proteins, the experimental data are scarce and are fragmented. For example, in [18], it is shown that chaperone DnaK of B. subtilis in vitro accomplishes refolding of denatured (5 M urea, 25°C) lactate dehy-
drogenase protein (LDH): after 50 min of incubation in a refolded medium containing ATP, phosphoenolpyruvate, NADH, pyruvate kinase, and DnaK, the percentage of active enzyme increased from 12 (spo
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