化学添加剂提高重组α-环糊精葡萄糖基转移酶酶制剂稳定性

Enhanced storage stability of recombinant enzyme preparation of α-CGTase from Paenibacillus macerans by chemical additives

摘要:

通过向重组α-环糊精葡萄糖基转移酶 (α-CGT 酶) 液中添加化学添加剂以提高其热稳定性及贮存稳定性。在不同温度下研究了添加剂对酶液的贮存稳定性影响，并用圆二色谱 (CD) 研究了CGT 酶在近紫外区和远紫外区蛋白质结构与热稳定性的变化关系。当单独加入各种添加剂在50 ℃水浴1 h 和室温放置108 d 后，发现含有20％甘油的酶液稳定性最好，与未加任何添加剂的对照酶液相比仍有91％和50％的酶活，对照酶液在50 ℃水浴1 h 后仅有小于10％的活性，室温放置108 d 后已经没有酶活。明胶、CaCl<

关键词:α-环糊精葡萄糖基转移酶; 化学添加剂; 热稳定性; 圆二色光谱

Abstract:

To enhance the thermostability and storage stability of α-cyclodextrin glycosyltransferase (α-CGTase), we added specific chemical additives into the preparation of α-CGTase, and studied the effect of additives on the storage stability of α-CGTase at different temperatures. Then we measured the protein structure of CGTase in the far UV (200?250 nm) and near UV (250?320 nm) ranges respectively by Circular dichroism (CD) spectra under high temperature and analyzed the relationship between thermostability and protein structure. The results indicated that the addition of selected additives (gelatin, glycerin, CaCl₂ and PEG400) enhanced the thermostability of α-CGTase dramatically. After 45 days, the preparation of α-CGTase still had 100% of the enzyme activity with different additives superimposed at the optimum concentration at 40 °C. The CD spectra of α-CGTase showed that glycerin could protect the secondary and the tertiary structure of the CGTase under high temperature and therefore the enzyme maintained its high activity. Chemical additives can improve the stability of α-CGTase significantly and they preserve the enzyme activity by protecting its secondary structure.

Key words:α-cyclodextrin glycosyltransferase; chemical additives; thermostability; circular dichroism