Optimization of the production and characterization of milk clotting enzymes by Bacillus subtilis natto

Abstract

Suitable medium for production of milk clotting enzyme (MCE) by Bacillus subtilis (natto) Takahashi in submerged liquid-state fermentation was screened, the nutrient factors affecting MCE production was optimized by response surface methodology. The MCE production by B. subtilis (natto) Takahashi was increased significantly by 428% in the optimal medium developed. The MCE was filtered and concentrated by ultrafiltration. The retentate after tandem filtration carried out with the combined membranes of MWCO 50kDa and 5 kDa showed two major bands between 25kDa and 30kDa on SDS-PAGE, and the MCA and MCA/PA improved significantly in comparison with those in the initial broth. The crude enzyme thus obtained showed MCA and MCA/PA ratio of 48,000 SU/g and 6,400, which are commensurate with those (MCA 26,667 SU/g and MCA/PA 6,667) of the commercial rennet. It had optimal pH and temperature at pH 6 and 60°C, and showed excellent pH and thermal stability.

Optimization of milk-clotting enzyme production by Bacillus amyloliquefaciens SP1 isolated from apple rhizosphere

“…However, the reason(s) for the decline remains to be investigated. Although similar results of fast MCE production by B. subtilis (natto), Takahashi and Thermomucor indicae were observed by Wu et al (2013) and Merheb-Dini et al (2012). However, optimal production of Nocardiopsis sp.…”

“…However, various animals, plants, and microbial proteases have been suggested as milk coagulants (Zhang et al 2011), but still attention has been focused on production of milk-clotting enzymes from microbial sources (Silveira et al 2005;Wu et al 2013) due to their rapid growth, relative inexpensive growth substrate, and diverse properties. Microbial rennet-like milkclotting enzymes are aspartic proteases that catalyze milk coagulation.…”

An Optimization Study of α-Amylase Production by <i>Aspergillus niger</i> ATCC 16404 Grown on Orange Waste Powder

“…The correlation and determination coefficients are shown in Table 3. The correlation coefficients high values R = 0.969, on one hand, and the determination coefficients R 2 (0.938) (R 2 is a global measure for the model quality), on the other hand, show that the selected model is representative of the production of α-amylase [13].…”

“…3) 5 repetitions of the central point (level 0). This framework makes it possible to study each factor at 5 levels ( Table 2), and to obtain the second degree equation the derivatives of which, compared to 0, make it possible to determine the extremums coded co-ordinates which will be, then, converted into actual values [8] [13]. The relationship between the coded levels and the real levels of the factors used in a central composite design is also displayed in Table 2.…”

“…α-amylase (E.C 3.2.1.1) is an extracellular enzyme which is widespread among higher plants, animals and production of milk clotting enzymes by Bacillus subtilis [13].…”