Isolation and Purification of Novel Thermostable Alkaline Lipase from Local Thermophilic Microorganism

Febriani¹, Rukman Hertadi¹, Prihardi Kahar², Akhmaloka¹ and Fida Madayanti¹*

¹Biochemistry Research Group, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Ganesha 10, Bandung Indonesia.

²Asian Center For Environmental Research, Meisei University, 2-1-1, Hodokubo, Hino-Shi Tokyo, 191-8506 Japan.

Corresponding Author E-mail:fida@chem.itb.ac.id,

ABSTRACT: A local thermophilic microorganism namely DMS-3 has been identified as thermostable alkaline lipase producing isolate. The bacterium was isolated from Domas hot spring, Tangkuban Perahu Mount, West Java. Based on gram staining test and Scanning Electron Microscopy, the microorganism showed a rod shape and a gram positive bacterium. The isolated showed maximum expression of lipase at pH 9 and 70oC. The lipase produced by DMS-3 isolate was purified using column chromatography of DEAE sepharose fast flow and Sephacryl S-200. Following Sephacryl separation the enzyme still showed specific activity at 54 times higher compared to that the wild type, with yield was about 4.89 %. Polyacryamide gel electrophoresis combining with zymogram analysis showed that there were 4 bands of protein exhibiting lipase activity. Further analysis by eluting of each band showed that the first, second, and third band revealed few bands similar of each other on denatured SDS-PAGE. Meanwhile the fourth band only showed single band. The data suggested that DMS-3 isolated expressed more than one type of lipases.

KEYWORDS: Thermostable alkaline lipase; microorganism thermophile; domas isolate.

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