Full-text resources of PSJD and other databases are now available in the new Library of Science.
Visit https://bibliotekanauki.pl


Preferences help
enabled [disable] Abstract
Number of results
2016 | 63 | 1 | 117-125

Article title

Cloning, expression, and biochemical characterization of a cold-active GDSL-esterase of a Pseudomonas sp. S9 isolated from Spitsbergen island soil


Title variants

Languages of publication



An estS9 gene, encoding an esterase of the psychrotolerant bacterium Pseudomonas sp. S9 was cloned and sequenced. The deduced sequence revealed a protein of 636 amino acid residues with a molecular mass of 69 kDa. Further amino acid sequence analysis revealed that the EstS9 enzyme contained a G-D-S-L motif centered at a catalytic serine, an N-terminal catalytic domain and a C-terminal autotransporter domain. Two recombinant E. coli strains for production of EstS9N (a two domain enzyme) and EstS9Δ (a one domain enzyme) proteins were constructed, respectively. Both recombinant proteins were successfully produced as inclusion bodies and then purified under denaturing conditions. However, because of the low enzymatic activity of the refolded EstS9Δ protein, only the EstS9N protein was further characterized. The purified and refolded EstS9N protein was active towards short-chain p-nitrophenyl esters (C2-C8), with optimal activity for the butyrate (C4) ester. With p-nitrophenyl butyrate as the substrate, the enzyme displayed optimal activity at 35°C and pH 9.0. Additionally, the EstS9N esterase retained ~90% of its activity from 25-40°C and ~40% of its activity at 10°C. Moreover, analysis of its kinetic parameters (Km, kcat, kcat/Km) toward p-nitrophenyl butyrate determined at 15°C and 25°C confirmed that the EstS9 enzyme is cold-adapted. To the best of our knowledge, EstS9 is the third characterized cold-active GDSL-esterase and the first one confirmed to contain an autotransporter domain characteristic for enzymes secreted by the type V secretion system.








Physical description




  • Department of Molecular Biotechnology and Microbiology, Gdańsk University of Technology, Gdańsk, Poland
  • Department of Molecular Biotechnology and Microbiology, Gdańsk University of Technology, Gdańsk, Poland
  • Department of Molecular Biotechnology and Microbiology, Gdańsk University of Technology, Gdańsk, Poland
  • Department of Molecular Biotechnology and Microbiology, Gdańsk University of Technology, Gdańsk, Poland
  • Department of Molecular Biotechnology and Microbiology, Gdańsk University of Technology, Gdańsk, Poland
  • Department of Molecular Biotechnology and Microbiology, Gdańsk University of Technology, Gdańsk, Poland


  • Abdul Salam J, Lakshmi V, Das D, Das N (2013) Biodegradation of lindane using a novel yeast strain, Rhodotorula sp. VITJzN03 isolated from agricultural soil. World J Microbiol Biotechnol 29: 475-87.
  • Akoh CC, Lee GC, Liaw YC, Huang TH, Shaw JF (2004) GDSL family of serine esterases/lipases. Prog Lipid Res 43: 534-552.
  • Arpigny JL, Jaeger KE (1999) Bacterial lipolytic enzymes: classification and properties. Biochem J 343: 177-83.
  • Berlemont R, Jacquin O, Delsaute M, La Salla M, Georis J, Verte F, Galleni M, Power P (2013) Novel cold-adapted esterase mhlip from an antarctic soil metagenome. Biology 2: 177-188.
  • Brault G, Shareck F, Hurtubise Y, Lepine F, Doucet N (2012) Isolation and characterization of EstC, a new cold-active esterase from Streptomyces coelicolor A3(2). PloS one 7: e32041.
  • Brzuszkiewicz A, Nowak E, Dauter Z, Dauter M, Cieslinski H, Dlugolecka A, Kur J (2009) Structure of EstA esterase from psychrotrophic Pseudoalteromonas sp 643A covalently inhibited by monoethylphosphonate. Acta Crystallogr Sect F Struct Biol Cryst Commun 65: 862-865.
  • Cavicchioli R, Charlton T, Ertan H, Mohd Omar S, Siddiqui KS, Williams TJ (2011) Biotechnological uses of enzymes from psychrophiles. Microb Biotechnol 4: 449-460.
  • Cieslinski H, Bialkowska AM, Dlugolecka A, Daroch M, Tkaczuk KL, Kalinowska H, Kur J, Turkiewicz M (2007) A cold-adapted esterase from psychrotrophic Pseudoalteromas sp strain 643A. Arch Microbiol 188: 27-36.
  • Dlugolecka A, Cieslinski H, Bruzdziak P, Gottfried K, Turkiewicz M, Kur J (2009) Purification and biochemical characteristic of a cold-active recombinant esterase from Pseudoalteromonas sp 643A under denaturing conditions. Pol J Microbiol 58: 211-218.
  • Dlugolecka A, Cieslinski H, Turkiewicz M, Bialkowska AM, Kur J (2008) Extracellular secretion of Pseudoalteromonas sp cold-adapted esterase in Escherichia coli in the presence of Pseudoalteromonas sp components of ABC transport system. Protein Expr Purif 62: 179-184.
  • Esteban-Torres M, Mancheno JM, de las Rivas B, Munoz R (2014) Characterization of a cold-active esterase from Lactobacillus plantarum suitable for food fermentations. J Agric Food Chem 62: 5126-5132.
  • Fojan P, Jonson PH, Petersen MT, Petersen SB (2000) What distinguishes an esterase from a lipase: a novel structural approach. Biochimie 82: 1033-1041.
  • Fu J, Leiros HK, de Pascale D, Johnson KA, Blencke HM, Landfald B (2013) Functional and structural studies of a novel cold-adapted esterase from an Arctic intertidal metagenomic library. Appl Microbiol Biotechnol 97: 3965-3978.
  • Fu C, Hu Y, Xie F, Guo H, Ashforth EJ, Polyak SW, Zhu B, Zhang L (2011) Molecular cloning and characterization of a new cold-active esterase from a deep-sea metagenomic library. Appl Microbiol Biotechnol 90: 961-970.
  • Heath C, Hu XP, Cary SC, Cowan D (2009) Identification of a novel alkaliphilic esterase active at low temperatures by screening a metagenomic library from antarctic desert soil. Appl Environ Microbiol 75: 4657-4659.
  • Henderson IR, Navarro-Garcia F, Desvaux M, Fernandez RC, Ala'Aldeen D (2004) Type V protein secretion pathway: the autotransporter story. Microbiol Mol Biol Rev 68: 692-744.
  • Henderson IR, Navarro-Garcia F, Nataro JP (1998) The great escape: structure and function of the autotransporter proteins. Trends Microbiol 6: 370-378.
  • Hu XP, Heath C, Taylor MP, Tuffin M, Cowan D (2012) A novel, extremely alkaliphilic and cold-active esterase from Antarctic desert soil. Extremophiles 16: 79-86.
  • Jang SH, Kim J, Kim J, Hong S, Lee C (2012) Genome sequence of cold-adapted Pseudomonas mandelii strain JR-1. J Bacteriol 194: 3263-3263.
  • Jiang X, Xu X, Huo Y, Wu Y, Zhu X, Zhang X, Wu M (2012) Identification and characterization of novel esterases from a deep-sea sediment metagenome Arch Microbiol 194: 207-214.
  • Jimenez DJ, Montana JS, Alvarez D, Baena S (2012) A novel cold active esterase derived from Colombian high Andean forest soil metagenome. World J Microbiol Biotechnol 28: 361-70.
  • Kang CH, Oh KH, Lee MH, Oh TK, Kim BH, Yoon J (2011) A novel family VII esterase with industrial potential from compost metagenomic library. Microb Cell Fact 10: 41.
  • Kim YO, Park IS, Nam BH, Kim DG, Jee YJ, Lee SJ, An CM (2014) A novel esterase from Paenibacillus sp. PBS-2 is a new member of the beta-lactamase belonging to the family VIII lipases/esterases. J Microbiol Biotechnol 24: 1260-1268.
  • Kulakova L, Galkin A, Nakayama T, Nishino T, Esaki N (2004) Cold-active esterase from Psychrobacter sp. Ant300: gene cloning, characterization, and the effects of Gly-->Pro substitution near the active site on its catalytic activity and stability. Biochim Biophys Acta 1696: 59-65.
  • Laemmli UK (1970) Cleavage of Structural Proteins during Assembly of Head of Bacteriophage-T4. Nature 227: 680-685.
  • Lemak S, Tchigvintsev A, Petit P, Flick R, Singer AU, Brown G, Evdokimova E, Egorova O, Gonzalez CF, Chernikova TN, Yakimov MM, Kube M, Reinhardt R, Golyshin PN, Savchenko A, Yakunin AF (2012) Structure and activity of the cold-active and anion-activated carboxyl esterase OLEI01171 from the oil-degrading marine bacterium Oleispira antarctica. Biochem J 445: 193-203.
  • Novototskaya-Vlasova K, Petrovskaya L, Yakimov S, Gilichinsky D (2012) Cloning, purification, and characterization of a cold-adapted esterase produced by Psychrobacter cryohalolentis K5T from Siberian cryopeg. Fems Microbiol Ecol 82: 367-375.
  • Reddy PV, Rao SS, Pratibha MS, Sailaja B, Kavya B, Manorama RR, Singh SM, Srinivas TN, Shivaji S (2009) Bacterial diversity and bioprospecting for cold-active enzymes from culturable bacteria associated with sediment from a melt water stream of Midtre Lovenbreen glacier, an Arctic glacier. Res Microbiol 160: 538-546.
  • Soror SH, Verma V, Rao R, Rasool S, Koul S, Qazi GN, Cullum J (2007) A cold-active esterase of Streptomyces coelicolor A3(2): from genome sequence to enzyme activity. J Ind Microbiol Biotechnol 34: 525-531.
  • Suzuki T, Nakayama T, Choo DW, Hirano Y, Kurihara T, Nishino T, Esaki N (2003) Cloning, heterologous expression, renaturation, and characterization of a cold-adapted esterase with unique primary structure from a psychrotroph Pseudomonas sp. strain B11-1. Protein Expr Purif 30: 171-178.
  • Suzuki T, Nakayama T, Kurihara T, Nishino T, Esaki N (2002) Primary structure and catalytic properties of a cold-active esterase from a psychrotroph, Acinetobacter sp. strain No. 6. isolated from Siberian soil. Biosci Biotechnol Biochem 66: 1682-1690.
  • Weisburg WG, Barns SM, Pelletier DA, Lane DJ (1991) 16S ribosomal DNA amplification for phylogenetic study. J Bacteriol 173: 697-703. .
  • Wierzbicka-Wos A, Bartasun P, Cieslinski H, Kur J (2013) Cloning and characterization of a novel cold-active glycoside hydrolase family 1 enzyme with beta-glucosidase, beta-fucosidase and beta-galactosidase activities. BMC Biotechnol 13: 22.
  • Wilhelm S, Tommassen J, Jaeger KE (1999) A novel lipolytic enzyme located in the outer membrane of Pseudomonas aeruginosa. J Bacteriol 181: 6977-6986.

Document Type

Publication order reference


YADDA identifier

JavaScript is turned off in your web browser. Turn it on to take full advantage of this site, then refresh the page.