A Novel Cyclodextrin Glycosyltransferase from Alkaliphilic Amphibacillus sp. NPST-10: Purification and Properties

International Journal of Molecular Sciences

Abdelnasser S. S. Ibrahim¹  Ali A. Al-Salamah¹  Mohamed A. El-Tayeb¹  Yahya B. El-Badawi¹ 

[1] Department of Botany and Microbiology, College of Science, King Saud University, Riyadh 11451, Kingdom of Saudi Arabia; E-Mails:

关键词: alkaliphiles;    soda lakes;    cyclodextrin glycosyltransferase;    Amphibacillus sp.;    purification;    16S rDNA;   

DOI  :  10.3390/ijms130810505

来源: mdpi

【 摘 要 】

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. K_m and V_max 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.

【 授权许可】

CC BY   
© 2012 by the authors; licensee Molecular Diversity Preservation International, Basel, Switzerland.