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A Novel Cyclodextrin Glycosyltransferase from Alkaliphilic Amphibacillus sp. NPST-10: Purification and Properties
Department of Botany and Microbiology, College of Science, King Saud University, Riyadh 11451, Kingdom of Saudi Arabia
Institute of Technical Microbiology, Hamburg University of Technology, Kasernenstrasse 12, 21073 Hamburg, Germany
* Author to whom correspondence should be addressed.
Received: 26 June 2012; in revised form: 5 August 2012 / Accepted: 10 August 2012 / Published: 22 August 2012
Abstract: Screening for cyclodextrin glycosyltransferase (CGTase)-producing alkaliphilic bacteria from samples collected from hyper saline soda lakes (Wadi Natrun Valley, Egypt), resulted in isolation of potent CGTase producing alkaliphilic bacterium, termed NPST-10. 16S rDNA sequence analysis identified the isolate as Amphibacillus sp. CGTase was purified to homogeneity up to 22.1 fold by starch adsorption and anion exchange chromatography with a yield of 44.7%. The purified enzyme was a monomeric protein with an estimated molecular weight of 92 kDa using SDS-PAGE. Catalytic activities of the enzyme were found to be 88.8 U mg−1 protein, 20.0 U mg−1 protein and 11.0 U mg−1 protein for cyclization, coupling and hydrolytic activities, respectively. The enzyme was stable over a wide pH range from pH 5.0 to 11.0, with a maximal activity at pH 8.0. CGTase exhibited activity over a wide temperature range from 45 °C to 70 °C, with maximal activity at 50 °C and was stable at 30 °C to 55 °C for at least 1 h. Thermal stability of the purified enzyme could be significantly improved in the presence of CaCl2. Km and Vmax values were estimated using soluble starch as a substrate to be 1.7 ± 0.15 mg/mL and 100 ± 2.0 μmol/min, respectively. CGTase was significantly inhibited in the presence of Co2+, Zn2+, Cu2+, Hg2+, Ba2+, Cd2+, and 2-mercaptoethanol. To the best of our knowledge, this is the first report of CGTase production by Amphibacillus sp. The achieved high conversion of insoluble raw corn starch into cyclodextrins (67.2%) with production of mainly β-CD (86.4%), makes Amphibacillus sp. NPST-10 desirable for the cyclodextrin production industry.
Keywords: alkaliphiles; soda lakes; cyclodextrin glycosyltransferase; Amphibacillus sp.; purification; 16S rDNA
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Ibrahim, A.S.S.; Al-Salamah, A.A.; El-Tayeb, M.A.; El-Badawi, Y.B.; Antranikian, G. A Novel Cyclodextrin Glycosyltransferase from Alkaliphilic Amphibacillus sp. NPST-10: Purification and Properties. Int. J. Mol. Sci. 2012, 13, 10505-10522.
Ibrahim ASS, Al-Salamah AA, El-Tayeb MA, El-Badawi YB, Antranikian G. A Novel Cyclodextrin Glycosyltransferase from Alkaliphilic Amphibacillus sp. NPST-10: Purification and Properties. International Journal of Molecular Sciences. 2012; 13(8):10505-10522.
Ibrahim, Abdelnasser S. S.; Al-Salamah, Ali A.; El-Tayeb, Mohamed A.; El-Badawi, Yahya B.; Antranikian, Garabed. 2012. "A Novel Cyclodextrin Glycosyltransferase from Alkaliphilic Amphibacillus sp. NPST-10: Purification and Properties." Int. J. Mol. Sci. 13, no. 8: 10505-10522.