论文部分内容阅读
从本实验室筛选得到的类球红杆菌(Rhodobacter sphaeroides)中,通过超声破碎、硫胺沉淀、DEAE-Sepha-dexA-25阴离子交换层析分离得到一种较纯的依赖NADPH的羰基还原酶.对其进行SDS-PAGE电泳分析,显示一条带,测得其相对分子量约为37kD.建立了羰基还原酶催化苯乙酮反应体系并对其进行优化,得出该酶催化苯乙酮的最适反应pH值为8,最适温度为37℃,在pH值为7～9之间比较稳定,其热稳定性较低.该酶对苯乙酮的米氏常数Km和最大反应速率Vmax分别为0.26mmol·L-1和2.4μmol·min-1·mg-1,最佳反应时间为24h·催化苯乙酮的主要产物为(S)-苯乙醇,其产率为58.5%,ee值可达到99%以上,是很有前景的生物催化剂之一.对酶催化与辅酶再生体系相结合进行了初步的研究,提出了氢化酶再生辅酶与菌绿素再生辅酶体系,为今后的酶催化工业生产奠定了基础.
Rhodobacter sphaeroides screened from our laboratory were separated by sonication, thiamine precipitation and DEAE-Sephadex-25 anion exchange chromatography to obtain a pure NADPH-dependent carbonyl reductase. SDS-PAGE electrophoresis analysis showed a band, measured the relative molecular weight of about 37kD.Carbonyl reductase catalyzed acetophenone reaction system and its optimization, the enzyme was catalyzed acetophenone optimum The reaction temperature is 8, the optimum temperature is 37 ℃, the pH value is between 7 and 9, and its thermal stability is lower.The Michaelis constant Km and maximum reaction rate Vmax of acetophenone are 0.26mmol·L-1 and 2.4μmol · min-1 · mg-1, the best reaction time was 24h. The main product of acetophenone was (S) -phenylethanol, the yield was 58.5% Reaching more than 99%, is one of the promising biocatalysts.Under the premise of the combination of enzyme-catalyzed and coenzyme regeneration system, the hydrogenase regeneration coenzyme and bacteriochlorin regeneration coenzyme system are proposed for the future enzyme catalytic industry Production laid the foundation.