Molecular characterization, gene expression analysis and biochemical properties of α-amylase from the disk abalone, Haliotis discus discus
Molecular characterization, gene expression analysis and biochemical properties of α-amylase from the disk abalone, Haliotis discus discus
复制标题
DOI：
10.1016/j.cbpb.2008.12.007
复制
发表时间：
2009-03-01
影响因子：
2.2
通讯作者：
Lee, Jehee
中科院分区：
文献类型：
--
作者：
Nikapitiya, Chamilani;Oh, Chulhong;Lee, Jehee
本研究报告了从盘鲍、七彩鲍 cDNA 文库中鉴定出的 α-淀粉酶的分子特征、克隆、表达和生化特征。 α-淀粉酶cDNA全长1650bp，编码511个氨基酸的多肽。成熟蛋白的预测 HdAmyl 分子量为 54 kDa，估计等电点 (pl) 为 8.3。 α-淀粉酶基因显示出其特征基序、催化位点、底物结合位点以及与其他已知α-淀粉酶种类的保守区域。纯化的重组 HdAmyl 对 1% 淀粉表现出相对较低的活性，为 0.1 U/mg 蛋白质。 HdAmyl 的最适温度和 pH 值分别为 50 摄氏度和 6.5。它还在很宽的温度和 pH 范围内表现出稳定性。组织特异性mRNA表达结果显示，HdAmyl仅在消化道和肝胰腺中表达，其中肝胰腺中含量最高。饥饿超过 8 周后，α-淀粉酶转录相对于基础水平显着下降。然而，饥饿后，mRNA 转录增加，并在喂养第 2 周恢复到正常水平，这表明盘鲍中 α-淀粉酶 mRNA 表达根据转录水平上食物利用率的变化而变化。 (C) 2008 Elsevier Inc. 保留所有权利。
The present study reports the molecular characterization, cloning, expression, and biochemical characterization of alpha-amylase identified from the disk abalone, Haliotis discus discus cDNA library. The full length of the alpha-amylase cDNA was 1650 bp, and it encoded a polypeptide of 511 amino acids. The predicted HdAmyl molecular mass of mature protein was 54 kDa and the estimated isoelectric point (pl) was 8.3. The alpha-amylase gene showed its characteristic motifs, catalytic sites, substrate binding sites and conserved regions with other known species of alpha-amylases. Purified recombinant HdAmyl exhibited a relatively low activity of 0.1 U/mg protein towards 1% starch. HdAmyl had an optimum temperature and pH of 50 degrees C and 6.5, respectively. It also demonstrated stability in a wide range of temperatures and pH. Tissue-specific mRNA expression results showed that HdAmyl is expressed only in the digestive tract and hepatopancreas, with the highest levels in the hepatopancreas. Over 8 weeks of starvation, alpha-amylase transcription was decreased significantly relative to basal levels. However, after starvation, mRNA transcription was increased and returned to normal level by the 2nd week of feeding, suggesting that the alpha-annylase mRNA expression changes according to variations in food availability at the transcriptional level in disk abalone. (C) 2008 Elsevier Inc. All rights reserved.