Part:BBa_K3077101

lip8 in Pseudomonas aeruginosa

This part, BBa_K3077101, is a cytoplasmic esterase encoded as lip8 in Pseudomonas aeruginosa.The genomic sequence is referenced from .[1] Lip8 gene consists of 1,173 nucleotides, encoding a protein of 391 amino acids[2].

Usage and Biology

Lip8 is a cytoplasmic esterase that catalyzes both the hydrolysis and synthesis of ester bonds. It is also a biocatalyst for interesterification, alcoholysis, acidolysis, and aminolysis.[2] It showed higher activities against short-chain fatty acid methyl esters compared with activities against various triacylglycerols and long-chain fatty acid methyl esters: Among triacylglycerols with different fatty acids, Lip8 was found to have the highest activity against triacetin (C2). While methyl acetate (C2), methyl propionate (C3), and methyl butyrate (C4) are the substrates with higher hydrolytic activity among different fatty acid methyl esters. Among four Natural oils, Lip8 showed highest activity against tung oil, whose principal fatty acid component is eleostearic acid (C18: 3)[2].

Family

Lip8 is a lipolytic enzyme belonging to family VIII, thus have serine residues in the -Ser-Xaa-Xaa-Lys- motif which acts as a catalytic nucleophile[2].

Conditions for working

No foldase is needed to assist the non-covalent folding of Lip8 when it is overexpressed. As it is a cytoplasmic gene, it is not accessible to its substrate unless the substrate passes through the outer membrane, or it leaks out of the cell in a significant amount. The optimum temperature and heat stability of Lip8 are lower than those of Lip3 and LST-03 lipase, two other lipolytic enzymes from the same strain[2]. It showed maximal activity at 30 ° C and pH 7, and is inactivated at more than 40 ° C[2].

The part is the gene of interest of the composite part BBa_K3077100.The ordered plasmid is transformed in E.coli and is verified through colony PCR with prefix forward and suffix reverse. We tested its catalytic activity as an esterase through the lipid hydrolysis assay.