Publications Repository - Gdańsk University of Technology

In this work, we present the construction of a metagenomic library in Escherichia coli using pUC19 vector and environmental DNA directly isolated from Antarctic topsoil and screened for lipolytic enzymes. The screening on agar supplemented with olive oil and rhodamine B revealed one clone with lipolytic activity (Lip1) out of 11,000 E. coli clones. This clone harbored a plasmid, pLip1, which has an insert of 4722 bp that has been completely sequenced from both directions. Further analysis of the insert showed three ORFs. The ORF2 encoded protein (Lip1) of 469 amino acids with 93% identity to the uncultured Pseudomonas sp. lipase LipJ03. Amino acid sequence comparison and phylogenetic analysis indicated that Lip1 lipase was closely related to family I subfamily 3. Furthermore, we present a three-dimensional model of the lipase Lip1 which was generated based on the two known structures of mesophilic lipases from Pseudomonas sp. MIS 38 (PML lipase, PDB; 2Z8X) and Serratia marcescens (SML lipase, PDB: 2QUB). Finally, we report the results of comparisons between the lipase Lip1 and mesophilic lipases and point out similarities and differences in the catalytic site and in other parts of the analyzed structures.

Authors

• dr hab. inż. Hubert Cieśliński link open in new tab ,
• Aneta Białkowska,
• Karolina Tkaczuk,
• Anna Długołęcka link open in new tab ,
• prof. dr hab. Józef Kur link open in new tab ,
• Marianna Turkiewicz