Enhancement of the Chaperone Activity of Alkyl Hydroperoxide Reductase C from Pseudomonas aeruginosa PAO1 Resulting from a Point-Specific Mutation Confers Heat Tolerance in Escherichia coli

• Published in: Molecules and cells Vol. 39; no. 8; pp. 594 - 602
• Main Authors: Lee, Jae Taek, Lee, Seung Sik, Mondal, Suvendu, Tripathi, Bhumi Nath, Kim, Siu, Lee, Keun Woo, Hong, Sung Hyun, Bai, Hyoung-Woo, Cho, Jae-Young, Chung, Byung Yeoup
• Format: Journal Article
• Language: English
• Published: United States Korean Society for Molecular and Cellular Biology 01.08.2016; 한국분자세포생물학회
• Subjects: Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Computational Biology; Computer Simulation; Escherichia coli - physiology; Heat-Shock Response - genetics; Hot Temperature - adverse effects; Molecular Chaperones - genetics; Molecular Chaperones - metabolism; Mutagenesis, Site-Directed; Peroxiredoxins - genetics; Peroxiredoxins - metabolism; Point Mutation - genetics; Protein Conformation; Protein Stability; Pseudomonas aeruginosa - physiology; Thermotolerance - genetics; Transgenes - genetics; 생물학; heat tolerance; alkyl hydroperoxide reductase subunit C; Pseudomonas aeruginosa; 2-Cys peroxiredoxin; chaperone
• ISSN: 1016-8478; 0219-1032
• DOI: 10.14348/molcells.2016.0042

Alkyl hydroperoxide reductase subunit C from Pseudomonas aeruginosa PAO1 (PaAhpC) is a member of the 2-Cys peroxiredoxin family. Here, we examined the peroxidase and molecular chaperone functions of PaAhpC using a site-directed mutagenesis approach by substitution of Ser and Thr residues with Cys at positions 78 and 105 located between two catalytic cysteines. Substitution of Ser with Cys at position 78 enhanced the chaperone activity of the mutant (S78C-PaAhpC) by approximately 9-fold compared with that of the wild-type protein (WT-PaAhpC). This increased activity may have been associated with the proportionate increase in the high-molecular-weight (HMW) fraction and enhanced hydrophobicity of S78C-PaAhpC. Homology modeling revealed that mutation of Ser(78) to Cys(78) resulted in a more compact decameric structure than that observed in WT-PaAhpC and decreased the atomic distance between the two neighboring sulfur atoms of Cys(78) in the dimer-dimer interface of S78C-PaAhpC, which could be responsible for the enhanced hydrophobic interaction at the dimer-dimer interface. Furthermore, complementation assays showed that S78C-PaAhpC exhibited greatly improved the heat tolerance, resulting in enhanced survival under thermal stress. Thus, addition of Cys at position 78 in PaAhpC modulated the functional shifting of this protein from a peroxidase to a chaperone.