摘要
建立了一种基于紫外显影法快速筛选烟酸羟基化酶产生菌的方法,用96孔板培养菌后,将菌液点样到滤纸上,利用6-羟基烟酸在紫外光照射(300nm)下显影的特性,可以快速、方便地筛选出烟酸羟基化酶产生菌.对筛选出的最高酶活菌株BKC4进行了研究,菌株为杆状,革兰氏染色呈阴性,通过16SrRNA序列分析鉴定为恶臭假单胞菌(Pseudomonas putida).利用BKC4菌株催化烟酸合成6-羟基烟酸,结果表明,BKC4菌株烟酸羟基化酶的最适催化条件为:底物浓度10g/L,温度37℃,pH6.5,摇床转速200r/min,酶在低于30℃及pH6.0～6.5范围内较稳定.
A rapid screening method for nicotine acid hydroxylase producing strain was established based on the ultraviolet development. The strain was cultured in 96-well microplate and a drop of broth was detected on the filter paper. The nicotine acid hydroxylase producing strain was screened fast and conveniently by the development of 6-hydroxynicotinic acid under ultraviolet illumination （300 nm）. Study on the strain BKC4 with the highest activity showed that it was rod, Gram negative bacterium, and identified as Pseudomonas putida by the analysis of 16S rRNA sequence. The strain of BKC4 was used to convert nicotinic acid to 6-hydroxynicotinic acid. The experimental result showed that the optimal catalytic conditions of BKC4 were 10 g/L of nicotinic acid, temperature 37 ℃,pH 6.5, 200 r/min of rotation speed, and the enzyme is stable at pH 6.5 and temperature below 30 ℃ .
出处
《过程工程学报》
CAS
CSCD
北大核心
2010年第3期576-581,共6页
The Chinese Journal of Process Engineering
基金
国家重点基础研究发展规划(973)基金资助项目(编号:2007CB714304)
关键词
烟酸羟基化酶
紫外显影
烟酸
6-羟基烟酸
nicotine acid hydroxylase
ultraviolet development
nicotinic acid
6-hydroxynicotinic acid
