Cloning, Purification, and Characterization of Recombinant Endo- β--1,4-D-Xylanase Of Bacillus sp. From Soil Termite Abdomen Safitri Eka(2), Hanifah(2), Previta(3), Sudarko(1), Tri Puspaningsih(3), Anak Agung Istri Ratnadewi(1,2)
(1) Departement of Chemistry, Faculty of Mathematics and Natural Sciences, University of Jember, Jalan Kalimantan 37, Jember 68121, Indonesia
(2) Graduate School of Biotechnology and PUI-BioTIn, University of Jember University of Jember, Jember 68121, Indonesia
(3) Department of Chemistry, Faculty of Sciences and Technology, Universitas Airlangga, Kampus C Mulyorejo, Surabaya 60115, Indonesia
Abstract
A novel endo beta 1,4 D Xylanase (xynBT) was isolated from Bacillus sp in termite abdominal soil. This enzyme is believed to produce XOS from cassava waste as a substrate. Since many agroindustrial wastes are a potentially valuable resource for industrial exploitation, this work aimed to cloning, overexpression, and endoxylanase enzyme characteristics. xynBT was successfully cloned in E. coli TOP 10 and expressed in E. coli BL21 (DE3) via pET30a. DNA sequence is containing an ORF of 801 bp nucleotide encoding a 267-aa. The molecular mass was 30 kDa. Based on the similarity (99%) of amino acid sequences, this enzyme is classified as a member of the glycoside hydrolase family 11. (rXynBT), which has been purified, shows an optimal pH and temperature of 4,6 and 40 degree celcius, respectively. Purified rXynBT was the most stable at pH 4 for up to 120 minutes pre-incubation time and had relative activity of 83%. This was stable at 30 until 40 degree celcius during 80 min pre incubation time with a relative activity of more than 50%. The presence of metal cations Kalium and Natrium on rXynBT increased its activity, while the metal cations Magnesium, Copper, Zinc and Iron on rXynBT were inhibitors.
