小麦麸皮抗冻蛋白的纯化及理化性质的研究

金周筠; 刘宝林

doi:10.13386/j.issn1002-0306.2015.20.025

小麦麸皮抗冻蛋白的纯化及理化性质的研究

Purification and physical-chemical characteristic of a novel antifreeze protein from wheat bran(wb AFP)

摘要

摘要: 本文纯化了一种小麦麸皮抗冻蛋白,并对其氨基酸组成、二级结构进行了分析。采用粗分离、饱和硫酸铵沉淀、离子交换色谱（DEAE-cellulose-26）、凝胶过滤色谱（Sephadex G75）四步对其进行纯化,纯化倍数为449.99倍,产率为4.17%,分子量为11.2 ku,热滞活性为0.169℃（5 mg/m L）。氨基酸组成结果表明:其甘氨酸含量较高（52.08 mol%）,亲水性好。将其与几种常见抗冻糖蛋白进行相关性分析,推断其是一种与冷诱导相关的蛋白。采用傅里叶红外光谱检测,获得其二级结构为:α-螺旋:25.96%²6.52%,β-折叠:21.69%²3.78%,β-转角:35.31%³7.61%,无规卷曲:13.69%¹5.08%。

Abstract: A novel antifreeze protein from wheat bran(wb AFP) was purified and the amino acid composition and the secondary structure were studied. This AFP was purified by four steps:crude extracted,ammonium sulfate precipitation, anion- exchange chromatography on a DEAE- cellulose- 26 column, and size- exclusion chromatography on a Sephadex G75 column. As a result,wb AFP was purified 449.99 fold,yielding a productivity of 4.17%. The molecular weight was 11.2 ku and the thermal hysteresis activity(THA) was 0.169 ℃ at a concentration of 5 mg/m L. The result of amino acid composition of wb AFP showed that it was glycine-rich(52.08 mol%) and had a relatively high hydrophilicity. Compared with several common antifreeze glycoproteins(AFGPs) it was inferred that wb AFP was related with cold-inducible protein. The secondary structure was determined by FT-IR spectra. The results were summarized as α-helix of 25.96%²6.52%,β-sheet of 21.69%²3.78%,β-turn of 35.31%³7.61% and random coil of 13.69%¹5.08%.