Biochemical Characterization of a GDSL-Motif Esterase from Bacillus sp. K91 with a New Putative Catalytic Mechanism = Biochemical Characterization of a GDSL-Motif Esterase from Bacillus sp. K91 with a New Putative Catalytic Mechanism

다국어 초록 (Multilingual Abstract)

The esterase gene Est8 from the thermophilic bacterium Bacillus sp. K91 was cloned and expressed in Escherichia coli. The monomeric enzyme exhibited a theoretical molecular mass of 24.5 kDa and an optimal activity around 50°C at pH 9.0. A model of Est8 was constructed using a hypothetical YxiM precursor structure (2O14_A) from Bacillus subtilis as template. The structure showed an α/β-hydrolase fold and indicated the presence of a typical catalytic triad consisting of Ser-11, Asp-182, and His-185, which were investigated by site directed replacements coupled with kinetic characterization. Asp-182 and His-185 residues were more critical than the Ser-11 residue in the catalytic activity of Est8. A comparison of the amino acid sequence showed that Est8 could be grouped into the GDSL family and further classified as an SGNH hydrolase. Est8 is a new member of the SGNH hydrolase subfamily and may employ a different catalytic mechanism.

참고문헌 (Reference)

1 Dalrymple BP, "Three Neocallimastix patriciarum esterases associated with the degradation of complex polysaccharides are members of a new family of hydrolases" 143 : 2605-2614, 1997

2 Arnold K, "The SWISSMODEL workspace : a web-based environment for protein structure homology modelling" 22 : 195-201, 2006

3 Yang C, "Surface display of MPH on Pseudomonas putida JS444 using ice nucleation protein and its application in detoxification of organophosphates" 99 : 30-37, 2008

4 Hedstrom L., "Serine protease mechanism and specificity" 102 : 4501-4524, 2002

5 Oh IS, "Secretome analysis reveals an Arabidopsis lipase involved in defense against Alternaria brassicicola" 17 : 2832-2847, 2005

6 Schwede T, "SWISSMODEL:An automated protein homology-modeling server" 31 : 3381-3385, 2003

7 Molgaard A, "Rhamnogalacturonan acetylesterase elucidates the structure and function of a new family of hydrolases" 8 : 373-383, 2000

8 Shaw JF, "Nucleotide sequence of a novel arylesterase gene from Vibro mimicus and characterization of the enzyme expressed in Escherichia coli" 298 : 675-680, 1994

9 Robertson DL, "Influence of active site and tyrosine modification on the secretion and activity of the Aeromonas hydrophila lipase/acyltransferase" 269 : 2146-2150, 1994

10 Pfeffer JM, "Identification of the first known inhibitors of O-acetylpeptidoglycan esterase : a potential new antibacterial target" 13 : 722-731, 2012

11 Weadge JT, "Identification and characterization of O-acetylpeptidoglycan esterase : a novel enzyme discovered in Neisseria gonorrhoeae" 45 : 839-851, 2006

12 Rubin B., "Grease pit chemistry exposed" 1 : 568-572, 1994

13 Akoh CC, "GDSL family of serine esterases/lipases" 43 : 534-552, 2004

14 Lee LC, "Functional role of catalytic triad and oxyanion hole-forming residues on enzyme activity of Escherichia coli thioesterase I/protease I/phospholipase L1" 397 : 69-76, 2006

15 Hong JK, "Function of a novel GDSL-type pepper lipase gene, CaGLIP1, in disease susceptibility and abiotic stress tolerance" 227 : 539-558, 2008

16 Hotta Y, "Extremely stable and versatile carboxylesterase from a hyperthermophilic archaeon" 68 : 3925-3931, 2002

17 Gouet P, "ESPript : analysis of multiple sequence alignments in PostScript" 15 : 305-308, 1999

18 Lo YC, "Crystal structure of Escherichia coli thioesterase I/protease I/lysophospholipase L1: consensus sequence blocks constitute the catalytic center of SGNH-hydrolases through a conserved hydrogen bond network" 330 : 539-551, 2003

19 Volokita M, "Combining comparative sequence and genomic data to ascertain phylogenetic relationships and explore the evolution of the large GDSL-lipase family in land plants" 28 : 551-565, 2011

20 Seung-Bum Kim, "Cloning and Characterization of Thermostable Esterase from Archaeoglobus fulgidus" 한국미생물학회 46 (46): 100-107, 2008

21 Thompson JD, "CLUSTAL W:improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice" 22 : 4673-4680, 1994

22 Zaide G, "Biochemical characterization and identification of catalytic residues in alpha-glucuronidase from Bacillus stearothermophilus T-6" 268 : 3006-3016, 2001

23 Jaeger KE, "Bacterial biocatalysts:molecular biology, three-dimensional structures, and biotechnological applications of lipases" 53 : 315-351, 1999

24 Laskowski RA, "AQUA and PROCHECK-NMR: programs for checking the quality of protein structures solved by NMR" 8 : 477-486, 1996

25 Upton C, "A new family of lipolytic enzymes?" 20 : 178-179, 1995

26 Morana A, "A carboxylesterase from the hyperthermophilic archaeon Sulfolobus solfataricus : cloning of the gene, characterization of the protein" 283 : 107-115, 2002