Skip to main content
SpringerLink
Log in

The cold-active Lip1 lipase from the Antarctic bacterium Pseudoalteromonas haloplanktis TAC125 is a member of a new bacterial lipolytic enzyme family

  • Original Paper
  • Published:
Extremophiles Aims and scope Submit manuscript

Abstract

The genome of the Antarctic bacterium Pseudoalteromonas haloplanktis TAC125 was searched for the presence of genes encoding ester-hydrolysing enzymes. Amongst the others, the gene PSHAa0051 coding for a putative secreted esterase/lipase was selected. The psychrophilic gene was cloned, functionally over-expressed in P. haloplanktis TAC125, and the recombinant product (after named PhTAC125 Lip1) was purified. PhTAC125 Lip1 was found to be associated to the outer membrane and exhibited higher enzymatic activity towards synthetic substrates with long acyl chains. A structural model was constructed using the structure of carboxylesterase Est30 from Geobacillus stearothermophilus as template. The model covered the central part of the protein with the exceptions of PhTAC125 Lip1 N- and C-terminal regions, where the psychrophilic protein displays extra-domains. The constructed model showed a typical α/β-hydrolase fold, and confirmed the presence of a canonical catalytic triad consisting of Ser, Asp and His. The sequence analysis showed that PhTAC125 Lip1 is distantly related to other lipolytic enzymes, but closely related to other putative psychrophilic esterases/lipases. The aligned proteins share common features, such as: (1) a conserved new active-site pentapeptide motif (LGG(F/L/Y)STG); (2) the likely extra-cytoplasmic localization, (3) the absence of a typical calcium-binding pocket, and (4) the absence of a canonical lid. These observations strongly suggest that aligned proteins constitute a novel lipase family, typical of psychrophilic marine γ-proteobacteria, and PhTAC125 Lip1 could be considered the first characterised member of this family.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  • Akoh CC, Lee GC, Liaw YC, Huang TH, Shaw JF (2004) GDSL family of serine esterases/lipases. Prog Lipid Res 43:534–552

    Article  PubMed  CAS  Google Scholar 

  • Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    Article  PubMed  CAS  Google Scholar 

  • Arpigny JL, Lamotte J, Gerday C (1997) Molecular adaptation to cold of an Antarctic bacterial lipase. J Mol Catal B-Enzym 3:29–35

    Article  CAS  Google Scholar 

  • Arpigny JL, Jaeger KE (1999) Bacterial lipolytic enzymes: classification and properties. Biochem J 343:177–183

    Article  PubMed  CAS  Google Scholar 

  • Aurilia V, Parracino A, Saviano M, Rossi M, D’Auria S (2007) The psychrophilic bacterium Pseudoalteromonas haloplanktis TAC125 possesses a gene coding for a cold-adapted feruloyl esterase activity that shares homology with esterase enzymes from γ-proteobacteria and yeast. Gene 397:51–57

    Article  PubMed  CAS  Google Scholar 

  • Baker NA, Sept D, Joseph S, Holst MJ, McCammon JA (2001) Electrostatics of nanosystems: application to microtubules and the ribosome. Proc Natl Acad Sci USA 98:10037–10041

    Article  PubMed  CAS  Google Scholar 

  • Berendsen HJC, van der Spoel D, van Drunen R (1995) GROMACS: a message-passing parallel molecular dynamics implementation. Comput Phys Commun 91:43–56

    Article  CAS  Google Scholar 

  • Bos MP, Robert V, Tommassen J (2007) Biogenesis of the Gram-negative bacterial outer membrane. Annu Rev Microbiol. doi:10.1146/annurev.micro.61.080706.093245

  • Choo DW, Kurihara T, Suzuki T, Soda K, Esaki N (1998) A cold adapted lipase of an Alaskan psychrotroph, Pseudomonas sp. strain B11–1: gene cloning and enzyme purification and characterization. Appl Environ Microbiol 64:486–491

    PubMed  CAS  Google Scholar 

  • Corpet F (1988) Multiple sequence alignment with hierarchical clustering. Nucleic Acids Res 16:10881–10890

    Article  PubMed  CAS  Google Scholar 

  • Cuff JA, Clamp ME, Siddiqui AS, Finlay M, Barton GJ (1998) Jpred: a consensus secondary structure prediction server. Bioinformatics 14:892–893

    Article  PubMed  CAS  Google Scholar 

  • D’Amico S, Collins T, Marx JC, Feller G, Gerday C (2006) Psychrophilic microorganisms: challenges for life. EMBO Rep 7:385–389

    Article  PubMed  CAS  Google Scholar 

  • Diaz P, Prim N, Pastor FIJ (1999) Direct fluorescence-based lipase activity assay. BioTechniques 27:696–677

    PubMed  CAS  Google Scholar 

  • Duilio A, Tutino ML, Marino G (2004) Recombinant protein production in Antarctic Gram-negative bacteria. Methods Mol Biol 267:225–237

    PubMed  CAS  Google Scholar 

  • Ewis HE, Abdelal AT, Lu CD (2004) Molecular cloning and characterization of two thermostable carboxyl esterases from Geobacillus stearothermophilus. Gene 329:187–195

    Article  PubMed  CAS  Google Scholar 

  • Fraternali F, Cavallo L (2002) Parameter optimized surfaces (POPS): analysis of key interactions and conformational changes in the ribosome. Nucleic Acids Res 30:2950–2960

    Article  PubMed  CAS  Google Scholar 

  • Hanahan D (1983) Studies on transformation of Escherichia coli with plasmids. J Mol Biol 166:557–580

    Article  PubMed  CAS  Google Scholar 

  • Handrick R, Reinhardt S, Focarete ML, Scandola M, Adamus G, Kowalczuk M, Jendrossek D (2001) A new type of thermoalkalophilic hydrolase of Paucimonas lemoignei with high specificity for amorphous polyesters of short chain-length hydroxylalkanoic acids. J Biol Chem 276:36215–36224

    Article  PubMed  CAS  Google Scholar 

  • Henikoff S, Henikoff JG (1992) Amino acid substitution matrices from protein blocks. Proc Natl Acad Sci USA 89:10915–10919

    Article  PubMed  CAS  Google Scholar 

  • Holmstrom C, James S, Neilan BA, White DC, Kjelleberg S (1998) Pseudoalteromonas tunicata sp. nov., a bacterium that produces antifouling agents. Int J Syst Bacteriol 48:1205–1212

    Article  PubMed  CAS  Google Scholar 

  • Iwai M, Tsujisaka Y, Fukumoto J (1964) Studies on lipase III. Effect of calcium ion on the action of the crystalline lipase from Aspergillus niger. J Gen Appl Microbiol 10:87–93

    Article  CAS  Google Scholar 

  • Jaeger KE, Ransac S, Dijkstra BW, Colson C, van Heuvel M, Misset O (1994) Bacterial lipase. FEMS Microbiol Rev 15:29–63

    Article  PubMed  CAS  Google Scholar 

  • Jaeger KE, Reetz M (1998) Microbial lipases from versatile tools for biotechnology. Trends Biotechnol 16:396–403

    Article  PubMed  CAS  Google Scholar 

  • Jaeger KE, Dijkstra BW, Reetz MT (1999) Bacterial biocatalysts: molecular biology, three dimensional structures, and biotechnological applications of lipases. Annu Rev Microbiol 53:315–351

    Article  PubMed  CAS  Google Scholar 

  • Jaeger KE, Eggert T (2002) Lipases for biotechnology. Curr Opin Biotechnol 13:390–397

    Article  PubMed  CAS  Google Scholar 

  • Jeong H, Yim JH, Lee C, Choi SH, Park YK, Yoon SH, Hur CG, Kang HY, Kim D, Lee HH, Park KH, Park SH, Park HS, Lee HK, Oh TK, Kim JF (2005) Genomic blueprint of Hahella chejuensis, a marine microbe producing an algicidal agent. Nucleic Acids Res 33:7066–7073

    Article  PubMed  CAS  Google Scholar 

  • Kwoun KH, Jung YJ, Choi WC, Ryu HS, Oh TK, Lee JK (2004) Sequence-based approach to finding functional lipases from microbial genome databases. FEMS Microbiol Lett 235:349–355

    Article  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  PubMed  CAS  Google Scholar 

  • Levisson M, van der Oost, Kengen SWM (2007) Characterization and structural modeling of a new type of thermostable esterase from Thermotoga maritima. FEBS 274:2832–2842

    Article  CAS  Google Scholar 

  • Liu P, Wang YF, Ewis HE, Abdelal AT, Lu CD, Harrison RW, Weber IT (2004) Covalent reaction intermediate revealed in crystal structure of the Geobacillus stearothermophilus carboxylesterase Est30. J Mol Biol 342:551–561

    Article  PubMed  CAS  Google Scholar 

  • Manco G, Di Gennaro S, De Rosa M, Rossi M (1994) Purification and characterization of a thermostable carboxylesterase from the thermoacidophilic eubacterium Bacillus acidocaldarius. Eur J Biochem 221:65–72

    Article  Google Scholar 

  • Margesin R, Schimner F (1994) Properties of cold adapted microorganisms and their role in biotechnology. J Biotechnol 33:1–4

    Article  CAS  Google Scholar 

  • Marti-Renom MA, Stuart A, Fiser A, Sánchez R, Melo F, Sali A (2000) Comparative protein structure modeling of genes and genomes. Annu Rev Biophys Biomol Struct 29:291–325

    Article  PubMed  CAS  Google Scholar 

  • Médigue C, Krin E, Pascal G, Barbe V, Bernsel A, Bertin PN, Cheung F, Cruveiller S, D’Amico S, Duilio A, Fang G, Feller G, Ho C, Mangenot S, Marino G, Nilsson J, Parrilli E., Rocha EPC, Rouy Z, Sekowska A, Tutino ML, Vallenet D, von Heijne G, Danchin A (2005) Coping with cold: the genome of the versatile marine Antarctica bacterium Pseudoalteromonas haloplanktis TAC125. Genome Res 15:1325–1335

    Article  PubMed  CAS  Google Scholar 

  • Mèthe BA, Nelson KE, Deming JW, Momen B, Melamud E, Zhang X, Moult J, Madupu R, Nelson WC, Dodson RJ, Brinkac LM, Daugherty SC, Durkin AS, DeBoy RT, Kolonay JF, Sullivan SA, Zhou L, Davidsen TM, Wu M, Huston AL, Lewis M, Weaver B, Weidman JF, Khouri H, Utterback TR, Feldblyum TV, Fraser CM (2005) The psychrophilic lifestyle as revealed by the genome sequence of Colwellia psychrerythraea 34H through genomic and proteomic analyses. Proc Natl Acad Sci USA 102:10913–10918

    Article  PubMed  CAS  Google Scholar 

  • Nardini M, Dijkstra BW (1999) Alpha/beta hydrolase fold enzymes: the family keeps growing. Curr Opin Struct Biol 9:732–737

    Article  PubMed  CAS  Google Scholar 

  • Papa R, Rippa V, Sannia G, Marino G, Duilio A (2007) An effective cold inducible expression system developed in Pseudoalteromonas haloplanktis TAC125. J Biotechnol 127:199–210

    Article  PubMed  CAS  Google Scholar 

  • Russell NJ (2000) Toward a molecular understanding of cold activity of enzymes from psychrophiles. Extremophiles 4:83–90

    Article  PubMed  CAS  Google Scholar 

  • Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • van Tilbeurgh H, Egloff MP, Martinez C, Rugani N, Verger R, Cambillau C (1993) Interfacial activation of the lipase-procolipase complex by mixed micelles revealed by X-ray crystallography. Nature 362:814–820

    Article  PubMed  Google Scholar 

  • Verger R (1997) Interfacial activation of lipases: facts and artifacts. Trends Biotechnol 15:32–38

    Article  CAS  Google Scholar 

  • Wilheilm S, Tomassen J, Jaeker KE (1999) A novel lipolytic enzyme located in the outer membrane of Pseudomonas aeruginosa. J Bacteriol 181:6977–6986

    Google Scholar 

Download references

Acknowledgments

This work was supported by grants of Ministero dell’Università e della Ricerca Scientifica (Progetti di Rilevante Interesse Nazionale 2006) and the National Programme of Antarctic Research 2004 (PNRA). Support the Regional Center of Competence (CRdC ATIBB, Regione Campania—Naples) is gratefully acknowledged.

Author information

Authors and Affiliations

  1. Institute of Protein Biochemistry, CNR, Via Pietro Castellino 111, 80131, Naples, Italy

    Donatella de Pascale & Guido di Prisco

  2. Department of Organic Chemistry and Biochemistry, University of Naples Federico II, Complesso Universitario, Monte Sant’Angelo, Via Cinthia 4, 80126, Naples, Italy

    Angela M. Cusano, Flavia Autore, Ermenegilda Parrilli, Gennaro Marino & M. Luisa Tutino

  3. School of Biotechnological Sciences, University of Naples Federico II, Complesso Universitario Monte Sant’Angelo, Via Cinthia 4, 80126, Naples, Italy

    Ermenegilda Parrilli, Gennaro Marino & M. Luisa Tutino

Authors
  1. Donatella de Pascale
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Angela M. Cusano
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Flavia Autore
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ermenegilda Parrilli
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Guido di Prisco
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Gennaro Marino
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. M. Luisa Tutino
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Donatella de Pascale.

Additional information

Communicated by K. Horikoshi.

D. de Pascale and A. M. Cusano equally contributed to the work.

Electronic supplementary material

Below is the link to the electronic supplementary material.

TABLE S1: Homologous sequences used as data set in phylogenetic analysis (TIF 82 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

de Pascale, D., Cusano, A.M., Autore, F. et al. The cold-active Lip1 lipase from the Antarctic bacterium Pseudoalteromonas haloplanktis TAC125 is a member of a new bacterial lipolytic enzyme family. Extremophiles 12, 311–323 (2008). https://doi.org/10.1007/s00792-008-0163-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00792-008-0163-9

Keywords

Search

Navigation