The metagenome-derived enzymes LipS and LipT increase the diversity of known lipases

PLoS One. 2012;7(10):e47665. doi: 10.1371/journal.pone.0047665. Epub 2012 Oct 24.


Triacylglycerol lipases (EC catalyze both hydrolysis and synthesis reactions with a broad spectrum of substrates rendering them especially suitable for many biotechnological applications. Most lipases used today originate from mesophilic organisms and are susceptible to thermal denaturation whereas only few possess high thermotolerance. Here, we report on the identification and characterization of two novel thermostable bacterial lipases identified by functional metagenomic screenings. Metagenomic libraries were constructed from enrichment cultures maintained at 65 to 75 °C and screened resulting in the identification of initially 10 clones with lipolytic activities. Subsequently, two ORFs were identified encoding lipases, LipS and LipT. Comparative sequence analyses suggested that both enzymes are members of novel lipase families. LipS is a 30.2 kDa protein and revealed a half-life of 48 h at 70 °C. The lipT gene encoded for a multimeric enzyme with a half-life of 3 h at 70 °C. LipS had an optimum temperature at 70 °C and LipT at 75 °C. Both enzymes catalyzed hydrolysis of long-chain (C(12) and C(14)) fatty acid esters and additionally hydrolyzed a number of industry-relevant substrates. LipS was highly specific for (R)-ibuprofen-phenyl ester with an enantiomeric excess (ee) of 99%. Furthermore, LipS was able to synthesize 1-propyl laurate and 1-tetradecyl myristate at 70 °C with rates similar to those of the lipase CalB from Candida antarctica. LipS represents the first example of a thermostable metagenome-derived lipase with significant synthesis activities. Its X-ray structure was solved with a resolution of 1.99 Å revealing an unusually compact lid structure.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alcohols / metabolism
  • Bacteria / chemistry
  • Bacteria / enzymology*
  • Bacteria / genetics
  • Bacteria / metabolism
  • Cloning, Molecular
  • Crystallography, X-Ray
  • DNA, Bacterial / genetics
  • Enzyme Stability
  • Esterification
  • Genome, Bacterial
  • Glycerides / metabolism
  • Lipase / chemistry*
  • Lipase / genetics
  • Lipase / metabolism*
  • Metagenome*
  • Metagenomics
  • Models, Molecular
  • Molecular Sequence Data
  • Nitrophenols / metabolism
  • Phylogeny
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Substrate Specificity
  • Up-Regulation


  • Alcohols
  • DNA, Bacterial
  • Glycerides
  • Nitrophenols
  • Recombinant Proteins
  • Lipase

Associated data

  • GENBANK/JQ028671
  • GENBANK/JQ028672
  • PDB/4FBL
  • PDB/4FBM

Grants and funding

This work was kindly funded by the German Federal Ministry of Education and Research (BMBF) cluster Biokatalyse2021 ( The research leading to the crystallization results has received funding from the European Community’s Seventh Framework Programme (FP7/2007–2013) under grant agreement n° 227764 (P-CUBE; K.-E. Jaeger and U. Krauss acknowledge support by the “Deutsche Forschungsgemeinschaft” (DFG) in frame of the research training group GK1166 “Biocatalysis in non-conventional media:” (BioNoCo; J. Pietruszka thanks the BMBF for funding the project “ExpresSys:” in the framework of the funding measure GenoMik ( The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.