title: Conserved tyrosine 182 residue in hyperthermophilic esterase EstE1 plays a critical role in stabilizing the active site.
abstract: An aromatic amino acid, Tyr or Trp, located in the esterase active site wall, is highly conserved, with hyperthermophilic esterases showing preference for Tyr and lower temperature esterases showing preference for Trp. In this study, we investigated the role of Tyr(182) in the active site wall of hyperthermophilic esterase EstE1. Mutation of Tyr to Phe or Ala had a moderate effect on EstE1 thermal stability. However, a small-to-large mutation such as Tyr to His or Trp had a devastating effect on thermal stability. All mutant EstE1 enzymes showed reduced catalytic rates and enhanced substrate affinities as compared with wild-type EstE1. Hydrogen bond formation involving Tyr(182) was unimportant for maintaining EstE1 thermal stability, as the EstE1 structure is already adapted to high temperatures via increased intramolecular interactions. However, removal of hydrogen bond from Tyr(182) significantly decreased EstE1 catalytic activity, suggesting its role in stabilization of the active site. These results suggest that Tyr is preferred over a similarly sized Phe residue or bulky His or Trp residue in the active site walls of hyperthermophilic esterases for stabilizing the active site and regulating catalytic activity at high temperatures.
authors.affiliation: Department of Biomedical Science and Center for Bio-Nanomaterials, Daegu University, Gyeongsan, 38453, South Korea.
authors.forename: Ngoc
authors.initials: N
authors.lastname: Truongvan
authors.affiliation: Department of Biological Sciences and Biotechnology, Hannam University, Daejeon, 34054, South Korea.
authors.forename: Hye-Shin
authors.initials: HS
authors.lastname: Chung
authors.affiliation: Department of Biomedical Science and Center for Bio-Nanomaterials, Daegu University, Gyeongsan, 38453, South Korea. shjang@daegu.ac.kr.
authors.forename: Sei-Heon
authors.initials: SH
authors.lastname: Jang
authors.affiliation: Department of Biomedical Science and Center for Bio-Nanomaterials, Daegu University, Gyeongsan, 38453, South Korea. leec@daegu.ac.kr.
authors.forename: ChangWoo
authors.initials: C
authors.lastname: Lee
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D017124:Conserved Sequence; D004795:Enzyme Stability; D004950:Esterases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006358:Hot Temperature; D014443:Tyrosine / Q000737:chemistry / Q000235:genetics
chemical_list: D019843:Archaeal Proteins; D001426:Bacterial Proteins; D014443:Tyrosine; D004950:Esterases
keywords: Active site; Esterase; Hyperthermophilic enzyme; Protein stability
references.citation: Int J Mol Sci. 2012;13(9):11643-65
references.pmid: 23109875
references.citation: Bioresour Technol. 2013 Apr;133:354-60
references.pmid: 23434813
references.citation: Appl Environ Microbiol. 2006 Apr;72(4):3021-5
references.pmid: 16598011
references.citation: J Mol Biol. 2001 Nov 30;314(3):507-18
references.pmid: 11846563
references.citation: Nucleic Acids Res. 2011 Jul;39(Web Server issue):W270-7
references.pmid: 21624888
references.citation: J Biol Chem. 2004 Jul 2;279(27):28202-8
references.pmid: 15100224
references.citation: Methods Mol Biol. 2008;426:299-318
references.pmid: 18542872
references.citation: FEBS J. 2012 Sep;279(17):3071-84
references.pmid: 22748144
references.citation: Extremophiles. 2015 Mar;19(2):235-47
references.pmid: 25472009
references.citation: Appl Biochem Biotechnol. 2013 Jan;169(1):29-40
references.pmid: 23117417
references.citation: Biochemistry. 2015 Mar 10;54(9):1841-8
references.pmid: 25711934
references.citation: PLoS One. 2011;6(5):e20506
references.pmid: 21637775
references.citation: Biochem Biophys Res Commun. 2014 Apr 18;446(4):1145-50
references.pmid: 24680822
references.citation: BMC Struct Biol. 2007 Jul 12;7:47
references.pmid: 17625021
references.citation: J Biol Chem. 2004 Feb 20;279(8):6815-23
references.pmid: 14617621
references.citation: Appl Environ Microbiol. 2005 Feb;71(2):817-25
references.pmid: 15691936
references.citation: Acta Crystallogr D Biol Crystallogr. 1999 Jul;55(Pt 7):1348-9
references.pmid: 10393304
references.citation: J Biol Chem. 2014 Aug 29;289(35):24499-510
references.pmid: 25043762
references.citation: Protein J. 2014 Aug;33(4):313-22
references.pmid: 24801996
references.citation: J Bacteriol. 2012 Jun;194(12):3263
references.pmid: 22628497
references.citation: Biochim Biophys Acta. 2014 Jun;1844(6):1076-82
references.pmid: 24667115
references.citation: Biotechnol Lett. 2012 Jun;34(6):1051-5
references.pmid: 22315100