Widhiastuty_2011_Int.J.Integr.Biol_11_8

A local thermophilic microorganism from Manuk hot spring, Kamojang, West Java, was isolated and cultivated. Homological analysis of the 16S rRNA sequences showed that the Manuk isolate is close to Geobacillus uzenensis strain X. These two microorganisms together with Geobacillus jurasicus develop a separate branch in phylogenetic tree. This group of Geobacillus strains has few unique 16S rRNA gene sequence or signature compared to that other Geobacillus strain. The lipase gene has been cloned and expressed in Escherichia coli, namely Lipase ITB 1.1. The gene contains 1248 bp coding for 416 amino acids sequence. The lipase is belong to sub-family 1.5 and close to Geobacillus stearothermophilus L1 lipase. Based on alignment analysis of its amino acid sequence, Lipase ITB 1.1 shows variation of amino acid residues with conserved residue of other lipase from sub-family 1.5. Pro395 residue in the conserved region of lipases was replaced by Leu in Lipase ITB 1.1. This residue together with Glu389, Asp394 and Gly308 residues were predicted to be involved in Ca-binding. Expression of the gene in Escherichia coli through pET-30a(+) expression vector showed that the protein was expressed as soluble protein with the expression level at around 30% of total protein.