Optimization and production of highly stable amylase isolated from Citrobacter portucalensis using low-cost agro-industrial wastes as substrates: Prospective therapeutics

Abstract

Starch hydrolyzing amylases are among the vital enzymes used in the food, textile, detergent, and pharmaceutical industries. The aim was to isolate amylase bacteria from agricultural soil, its extraction and subsequent purification. Within the scope of this work, using a 1% starch agar medium, potent thermostable bacterial amylase (A-4) was isolated from agricultural soil samples. Molecular analysis revealed the amylase-producing bacterial strain (A-4) to be Citrobacter portucalensis. We used agro-industrial-based wastes (wheat bran, potato peel, and sugarcane bagasse) in amylase production. Culture condition optimization showed that high amylase activity of 4.96 U/mL was obtained at temperature 55 °C and pH 5.0 after 72 h of incubation. The best-suited substrate was nitrogen, and carbon sources were wheat bran, yeast, and lactose for enzyme production. In addition, metal ions Ca+2 and Mg+2 showed a net positive effect on the rate of the enzymatic reaction, and Al+3 and Co+2 inhibited the activity. Protein purification results demonstrated a 4-fold increase with 13% yield and specific activity of 40 U/mL/min. SDS-PAGE indicated the molecular weight of partially purified protein as ∼56 kDa. The stain removal efficiency of the purified enzyme showed considerable results. The finding revealed the use of low-cost and readily available substrates, making C. portucalensis (A-4) a promising candidate for detergent and pharmaceutical industries.