Doctorate Program in Food Science and Technology, Faculty of Technology, University of Santiago of Chile, Chile.
Department of Biology, Faculty of Chemistry and Biology, University of Santiago of Chile, Chile.
Food Chem. 2018 Jul 15;254:87-94. doi: 10.1016/j.foodchem.2018.01.179. Epub 2018 Feb 2.
Myrosinase is a glycosylated enzyme present in the Brassicaceae family that catalyzes the hydrolysis of glucoraphanin to yield sulforaphane, recognized as a health-promoting compound found in cruciferous foods. Broccoli myrosinase has been poorly characterized. In this work, the enzyme was purified from broccoli florets and its kinetic behaviour was analyzed. The cDNA of broccoli myrosinase was isolated and sequenced to obtain the amino acids sequence of the enzyme. A three-dimensional structural model of a broccoli myrosinase subunit was built and used to perform molecular docking simulations with glucoraphanin and other glucosinolates. Kinetic data were adjusted to the Two-Binding Sites Model that describes substrate inhibition, obtaining R higher than 97%. The docking simulations confirmed the existence of two substrate-binding sites in the monomer, and allowed identifying the residues that interact with the substrate in each site. Our findings will help to design strategies to better exploit the health-promoting properties of broccoli.
黑芥子酶是存在于十字花科植物中的一种糖基化酶，能够催化葡萄糖硫苷水解生成具有促进健康作用的化合物——萝卜硫素，这种化合物存在于十字花科蔬菜中。西兰花黑芥子酶的特性尚未得到充分研究。本研究从西兰花花朵中纯化了该酶，并分析了其动力学行为。克隆并测序了西兰花黑芥子酶的 cDNA，获得了该酶的氨基酸序列。构建了西兰花黑芥子酶亚基的三维结构模型，并利用该模型进行了与葡萄糖硫苷和其他硫代葡萄糖苷的分子对接模拟。将动力学数据拟合到描述底物抑制的双结合位点模型中，得到的 R ² 值高于 97%。对接模拟证实了单体中存在两个底物结合位点，并确定了每个结合位点与底物相互作用的残基。我们的研究结果将有助于设计策略，更好地利用西兰花的健康促进特性。
