Purification and partial characterization of L-asparaginase enzyme produced by newly isolated Bacillus sp

Jimat, Dzun Noraini and Mohamed, Intan Baizura Firda and Azmi, Azlin Suhaida and Jamal, Parveen (2017) Purification and partial characterization of L-asparaginase enzyme produced by newly isolated Bacillus sp. IIUM Engineering Journal, 18 (2). ISSN 1511-788X E-ISSN 2289-7860

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Abstract

A new bacterial producing L-asparaginase was successfully isolated from Sungai Klah Hot Spring, Perak, Malaysia and identified as Bacillus sp. It was the best L-asparaginase producer as compared to other isolates. Production of L-asparaginase from the microbial strain was carried out under liquid fermentation. The crude enzyme was then centrifuged and precipitated with ammonium sulfate before being further purified by chromatographic method. HiTrap DEAE-Sepharose Fast Flow ion exchange chromatography followed by separation on Superose 12 gel filtration were used to obtain pure enzyme. The purified enzyme showed 10.11 U/mg of specific activity, 50.07% yield with 2.21 fold purification. The purified enzyme was found to be dimer in form, with a molecular weight of 65 kDa as estimated by SDS-PAGE. The maximum activity of the purified L-asparaginase was observed at pH 9 and temperature of 60 oC. Kajian penyelidikan ini telah berjaya menghasilkan enzim L-aspraginase daripada bakteria-bakteria baru yang diambil dari Kolam Air Panas, Sungai Klah, Perak, Malaysia dan ia dikenal pasti sebagai Bacillus sp. Teknik isolasi bakteria ini adalah teknik terbaik dalam menghasilkan enzim L-asparaginase berbanding teknik-teknik lain. Penghasilan enzim ini dibuat menerusi proses fermentasi. Kultur bakteria yang diperolehi diempar dan diikuti presipitasi menggunakan ammonium sulfat sebelum proses penulenan seterusnya dilakukan menggunakan kaedah kromatografi. Kolum penukaran ion jenis HiTrap DEAE-Sepharose Fast Flow diikuti dengan pemisahan oleh gel Superose 12 telah digunakan untuk mendapatkan enzim L-asparaginase yang tulen. Hasil kajian mendapati enzim tulen yang diperolehi mempunyai aktiviti spesifik sebanyak 10.11 U/mg daripada 50.07% enzim yang dihasilkan dan berjaya mencapai 2.21 kali ganda tulen dari enzim tanpa proses penulenan. Enzim tulen yang diperolehi didapati dalam bentuk “dimer“ dengan berat molekular sebanyak 65 kDa yang ditentukan menerusi SDS-PAGE. Enzim ini juga menunjukkan aktiviti yang tinggi pada pH 9 dan suhu 60 oC.

Item Type:	Article (Journal)
Additional Information:	3891/59636
Uncontrolled Keywords:	Bacillus sp.; L-asparaginase; ion exchange chromatography; gel filtration chromatography; partial characterization
Subjects:	Q Science > QR Microbiology T Technology > TP Chemical technology > TP248.13 Biotechnology
Kulliyyahs/Centres/Divisions/Institutes (Can select more than one option. Press CONTROL button):	Kulliyyah of Engineering > Department of Biotechnology Engineering
Depositing User:	Dr Dzun Noraini Jimat
Date Deposited:	24 Nov 2017 15:38
Last Modified:	20 Mar 2018 13:16
URI:	http://irep.iium.edu.my/id/eprint/59636

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