高去酰胺蛋白酶产生菌的诱变选育及产酶条件优化

以地衣芽孢杆菌2709为出发菌株,采用氮离子注入技术,在最佳注入剂量为1.5×1015ions/cm2的条件下进行诱变处理,筛选得到一株去酰胺度高达48.99%的突变株SDL-9,去酰胺度提高了85.57%,而肽键水解度相对较弱。并对SDL-9的产酶条件进行优化,结果表明:最佳碳源为质量浓度为2.0g/100mL的玉米粉,最佳氮源为质量浓度3.0g/100mL的大豆蛋白,质量浓度为0.04g/100mL Fe3+对产酶具有明显的促进作用,质量浓度为0.9g/100mL谷氨酰胺对产酶有明显的诱导作用,最佳起始pH值为7.0,最佳发酵温度35℃,在此条件下进行发酵产酶,测得去酰胺度为57.1%,肽键水解度为13.2%。

Breeding of Strain Producing Protease with High Deamidation and the Optimization of Fermentation Conditions

The original strain Bacillus licheniformis 2709 was mutagenized by means of nitrogen ion implantation at the optimal implantation dose of 1.5 × 1015 ions/cm2.As a result,a mutant strain with a deamidation capacity of 48.99%(increased by 85.57% compared with the original strain) but a relatively weak ability to cleave peptide bonds named as SDL-9 was obtained.The orthogonal array design method was used for the optimization of fermentation conditions for production of alkaline protease with high deamidation capacity by SDL-9.The optimal fermentation conditions were 2 g/100 mL corn meal as the carbon source,3 g/100 mL soybean protein as the nitrogen source,0.04 g/100 mL Fe3+ with distinct promoting effect on enzyme production,0.9 g/100 mL glutamine that could notably induce enzyme production,initial pH 7.0,and temperature 35 ℃.Under these conditions,the deamidation capacity and hydrolysis degree of peptide bonds were 57.1% and 13.2%,respectively.