Skip to main content
Log in

Cloning and expression of a novel lipase gene from Bacillus sphaericus 205y

  • Original Paper
  • Published:
Molecular Genetics and Genomics Aims and scope Submit manuscript

Abstract

A Bacillus sphaericus strain (205y) that produces an organic solvent-tolerant lipase was isolated in Port Dickson, Malaysia. The gene for the lipase was recovered from a genomic library and sequenced. Phylogenetic analysis was performed based on an alignment of thirteen microbial lipase sequences obtained from the NCBI database. The analysis suggested that the B. sphaericus lipase gene is a novel gene, as it is distinct from other lipase genes in Families I.4 and I.5 reported so far. Expression in Escherichia coli under the control of the lacZ promoter resulted in an eight-fold increase in enzyme activity after a 3-h induction with 1 mM IPTG. The crude enzyme thus obtained showed a slight (10%) enhancement in activity after a 30-min incubation in 25% (v/v) n -hexane at 37°C, and retained 90% of its activity after a similar period in 25% (v/v) p -xylene.

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

Access this article

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

Instant access to the full article PDF.

Fig. 1.
Fig. 2.
Fig. 3.
Fig. 4.
Fig. 5.
Fig. 6.
Fig. 7.
Fig. 8.

Similar content being viewed by others

References

  • Aono R, Itoh M, Inoue A, Horikoshi K (1992) Isolation of novel toluene-tolerant strain of Pseudomonas aeruginosa. Biosci Biotechnol Biochem 56:145–146

    CAS  Google Scholar 

  • Basri M, Heng AC, Razak CNA, Yunus W, Ahmad M, Rahman RNA, Ampon K, Salleh AB (1997) Alcoholysis of palm oil mid-fraction by lipase from Rhizopus rhizopodiformis. J Am Oil Chem Soc 74:113–116

    CAS  Google Scholar 

  • Brown TA (1990) Gene cloning: an introduction (2nd edn). Chapman and Hall, London

    Google Scholar 

  • Dartois V, Coppee JY, Colson C, Baulard A (1994) Genetic analysis and overexpression of lipolytic activity in Bacillus subtilis. Appl Environ Microbiol 60:1670–1673

    PubMed  Google Scholar 

  • Gierach LM (1989) Signal sequences. Biochemistry 28:923–930

    PubMed  Google Scholar 

  • Guruprasad K, Reddy BVB, Pandit MW (1990) Instability index. Prot Eng 4:155–161

    CAS  Google Scholar 

  • Henrissat B, Bairoch A (1993) New families in the classification of glycosyl hydrolases based on amino acid sequence. J Biochem 293:781–788

    CAS  Google Scholar 

  • Ikai A (1980) Aliphatic index. J Biochem 88:1895–1898

    CAS  PubMed  Google Scholar 

  • Inoue A, Yamamoto M, Horikoshi K (1991) Pseudomonas putida which can grow in the presence of toluene. Appl Environ Microbiol 57:1560–1562

    CAS  Google Scholar 

  • Kouker G, Jaeger KE (1987) Specific and sensitive assay for bacterial lipases. Appl Environ Microbiol 53:211–213

    CAS  PubMed  Google Scholar 

  • Kwon D Y, Rhee JS (1986) A simple and rapid colorimetric method for determination of free fatty acids for lipase assays. J Am Oil Chem Soc 63:29–32

    Google Scholar 

  • Laane C, Boeren S, Vos K, Veeder C (1987) Rules for optimization of biocatalysis in organic solvents. Biotech Bioeng 30:81–87

    CAS  Google Scholar 

  • Nawani N, Dosanjh NS, Kaur J (1998) A novel thermostable lipase from a thermophilic Bacillus sp.: characterization and esterification studies. Biotechnol Lett 20:997–1000

    Article  CAS  Google Scholar 

  • Ogino H, Ishikawa H (2001) Enzymes which are stable in the presence of organic solvents. J Biosci Bioeng 91:109–116

    Article  CAS  Google Scholar 

  • Ogino H, Miyamoto K, Ishikawa H (1994) Organic-solvent-tolerant bacterium which secretes organic-solvent-stable lipolytic enzyme. Appl Environ Microbiol 60:3884–3886

    CAS  Google Scholar 

  • Ogino H, Watanabe F, Yamada M, Nakagawa S, Hirose T, Noguchi A, Yasuda M, Ishikawa H (1999) Purification and characterization of organic solvent-stable protease from organic solvent-tolerant Pseudomonas aeruginosa PST-01. J Biosci Bioeng 87:61–68

    CAS  Google Scholar 

  • Palsan SS (1998) MS Thesis, Universiti Putra Malaysia, Serdang

  • Philips A, Pretorius GHJ, Rensburg HG (1995) Molecular characterization of a Galactomyces geotrichum lipase, another member of the cholinesterase/lipase family. Biochim Biophys Acta 1252:305–311

    PubMed  Google Scholar 

  • Samad MYA, Salleh AB, Razak CNA, Ampon K, Yunus WMZW, Basri M (1990) A lipase from a newly isolated thermophilic Rhizopus rhizopodiformis. World J Microb Biotechnol 6:390–394

    CAS  Google Scholar 

  • Sambrook J, Frisch ER, Maniatis T (1989) Molecular cloning: a laboratory manual (2nd edn). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.

    Google Scholar 

  • Schmidt-Dannert, Schmid RD, Menge U, Stocklein W, Sztajer H (1994) Screening, purification and properties of a thermophilic lipase from Bacillus thermocatenulatus. Biochim Biophys Acta 1214:43–53

    CAS  PubMed  Google Scholar 

  • Sugihara A, Shimada Y, Tominaga Y (1990) Separation and characterization of two molecular forms of Geotrichum candidum lipase. J Biochem 107:426–430

    CAS  PubMed  Google Scholar 

  • Sugihara A, Tani T, Tominaga Y (1991) Purification and characterization of a novel thermostable lipase from Bacillus sp. J Biochem 109:211–216

    CAS  PubMed  Google Scholar 

  • Sullivan ER, Leahy JG, Colwell RR (1999) Cloning and sequence analysis of the lipase and lipase chaperone-encoding genes from Acinetobacter calcoaceticus RAG-1, and redefinition of a Proteobacterial lipase family and an analogous lipase chaperone family. Gene 230:277–285

    Article  CAS  PubMed  Google Scholar 

  • Wohlfarth S, Winkler UK (1988) Chromosomal mapping and cloning of the lipase gene of Pseudomonas aeruginosa. J Gen Microbiol 134:433–440

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This research project was supported by IRPA Grant No. 51525 (09-02-04-0122)

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to R. N. Z. A. Rahman.

Additional information

Communicated by H. Ikeda

Rights and permissions

Reprints and permissions

About this article

Cite this article

Rahman, R.N.Z.A., Chin, J.H., Salleh, A.B. et al. Cloning and expression of a novel lipase gene from Bacillus sphaericus 205y. Mol Gen Genomics 269, 252–260 (2003). https://doi.org/10.1007/s00438-003-0831-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00438-003-0831-5

Keywords

Navigation