Murakami Masahito, Saito Makoto, Yokobori Hirokazu, Nishimura Katsushi, Tanigawa Minoru, Nagata Yoko
Department of Materials and Applied Chemistry, College of Science and Technology, Nihon University, Chiyoda-Ward, Tokyo, 101-8308, Japan.
Extremophiles. 2018 Jan;22(1):99-107. doi: 10.1007/s00792-017-0980-9. Epub 2017 Nov 9.
Pyrobaculum islandicum is a hyperthermophilic archaeon that grows optimally at 95-100 °C. In the previous study, we extensively purified a serine racemase from this organism and cloned the gene for overexpression in Escherichia coli (Ohnishi et al. 2008). This enzyme also exhibits highly thermostable L-serine/L-threonine dehydratase activity. In the present study, we aimed to elucidate the molecular mechanisms underlying the high thermostability of this enzyme. A recombinant variant of this enzyme, PiSRvt, constructed by truncating the C-terminal 72 amino acids, was compared with the native enzyme, PiSR. The dehydratase activity of PiSR and PiSRvt was found to owe to a homotrimer and a monomer, respectively, that demonstrated high and moderate thermostability, respectively. These observations reveal that the C-terminal region contributes to monomer trimerization that provides the extreme thermostability.
冰岛嗜火栖热菌是一种嗜热古菌，其最佳生长温度为95-100°C。在之前的研究中，我们从这种生物体中大量纯化了一种丝氨酸消旋酶，并克隆了该基因以便在大肠杆菌中进行过表达（大西等人，2008年）。这种酶还表现出高度耐热的L-丝氨酸/L-苏氨酸脱水酶活性。在本研究中，我们旨在阐明这种酶具有高耐热性的分子机制。通过截短C末端的72个氨基酸构建的该酶的重组变体PiSRvt，与天然酶PiSR进行了比较。发现PiSR和PiSRvt的脱水酶活性分别归因于同三聚体和单体，它们分别表现出高耐热性和中等耐热性。这些观察结果表明，C末端区域有助于单体三聚化，从而提供极高的耐热性。
