Production, Purification and Characterization of Alfa-amylase produced by Bacteria from Pharaonic Lake

Hazaa, Mahmoud M.; Sabae, Shawky Z.; T, Areej Ibrahim F.; Eldourghamy, Ayman S.; Sayed, H.

doi:10.21608/jbes.2018.370465

Production, Purification and Characterization of Alfa-amylase produced by Bacteria from Pharaonic Lake

Document Type : Original Article

Authors

¹ Department of plant, Faculty of science, Benha University, Benha, Egypt

² National Institute of oceanography and fisheries, Egypt

³ National Water Research Centre, El-Qanater El- Khayreia, Egypt

⁴ 4 Env. Biotech. Dept. Gen. Eng. Biotech. Res. Ins. University of Sadat City

10.21608/jbes.2018.370465

Abstract

In the present study thirty bacteria isolates collected from Pharaonic lake, Menoufiya Governorate, Egypt. Out
of which 12 isolates that were α-amylase producer. Isolate No. 1 was selected as the best enzyme producer,
identified as Alloiococcus otitis and was used for further studies. Attempts were made to optimize the cultural
condition for getting high yields of enzyme. The optimum level of pH for enzyme production was 8.0 on Bennett
medium after 8 hours. Among carbon sources, maltose supported maximum production of α-amylase followed by
lactose, sucrose, starch and manitol. Urea was the best nitrogen source for enzyme production, followed by L- asparagine. The purified fraction detected by the appearance of a single band corresponding to 38 k Da in SDS- PAGE, and the enzyme was purified 67.0-fold with amylase units 225 ml and specific activity of 234.4 U/mg.
The pure enzyme was stable at all degrees of temperatures (20–70ºC). The maximum activity was at pH of 9.0.
Starch was the best substrate. CaCl2, MgCl2 and KCl were the best activators. There is a continuous decreasing in
the activity due to the increase of SDS and H2O2 concentrations. Enzyme was stable in presence of butanol, n- hexane and toluene.

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