Isolation, expression, and characterization of raw starch degrading α-amylase from a marine lake Bacillus megaterium NL3

Sofi Siti Shofiyah; Dewi Yuliani; Nurul Widya; Fean D Sarian; Fernita Puspasari; Ocky Karna Radjasa; Ihsanawati; Dessy Natalia

doi:10.1016/j.heliyon.2020.e05796

Isolation, expression, and characterization of raw starch degrading α-amylase from a marine lake Bacillus megaterium NL3

Heliyon. 2020 Dec 26;6(12):e05796. doi: 10.1016/j.heliyon.2020.e05796. eCollection 2020 Dec.

Authors

Sofi Siti Shofiyah^{1

2}, Dewi Yuliani¹, Nurul Widya¹, Fean D Sarian¹, Fernita Puspasari¹, Ocky Karna Radjasa^{3

4}, Ihsanawati¹, Dessy Natalia^{1

5}

Affiliations

¹ Biochemistry Research Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesha No. 10, Bandung, 40132, Indonesia.
² Marine Science Program Study, Faculty of Marine Science, OSO University, Pontianak, 78113, Indonesia.
³ Center for Tropical Coastal and Marine Studies, Diponegoro University, Widya Puraya, Semarang, 50275, Indonesia.
⁴ Indonesian Institute of Sciences, Gatot Subroto 10, Jakarta, Indonesia.
⁵ University Center of Excellence for Nutraceuticals, Biosciences and Biotechnology Research Center, Bandung, Indonesia.

Abstract

A land-locked marine lake Kakaban with its significant ecological paramaters provides a unique habitat for bacteria with novel biotechnology potential that uses a diverse array of catalytic agents, including α-amylase. Aiming at the isolation of raw starch degrading α-amylase from marine biodiversity, a gene encoding BmaN2 from a sea anemone associated bacterium Bacillus megaterium NL3 was cloned and expressed in Escherichia coli ArcticExpress (DE3). It comprises an open reading frame of 1,563 nucleotides encoding BmaN2 of 520 amino acids and belongs to the glycoside hydrolase family 13 subfamily 36 (GH13_36). This α-amylase has a maximum activity at pH 6.0 and 60 °C with a specific activity of 28.7 U mg^-1. The BmaN2 activity is enhanced strongly by Ca²⁺ but inhibited by EDTA. BmaN2 also exhibits high catalytic efficiency on soluble starch with k _cat /K _M value of 14.1 mL mg^-1 s^-1. Despite no additional starch-binding domain, BmaN2 is able to hydrolyze various raw starches, such as wheat, corn, cassava, potato, rice, sago, and canna, in which granular wheat is the preferred substrate for BmaN2. These characteristics indicate that BmaN2 is a promising raw starch degrading enzyme within the subfamily GH13_36.

Keywords: Bacillus megaterium NL3; Marine microorganism; Raw starch degrading enzyme; α-amylase.