College of Life Sciences, Ritsumeikan University, Kusatsu, Shiga, 525-8577, Japan.
Ritsumeikan Global Innovation Research Organization, Ritsumeikan University, Kusatsu, Shiga, 525-8577, Japan.
Carbohydr Res. 2019 May 15;477:20-25. doi: 10.1016/j.carres.2019.03.011. Epub 2019 Mar 23.
UDP-apiose, a donor substrate of apiosyltransferases, is labile because of its intramolecular self-cyclization ability, resulting in the formation of apiofuranosyl-1,2-cyclic phosphate. Therefore, stabilization of UDP-apiose is indispensable for its availability and identifying and characterizing the apiosyltransferases involved in the biosynthesis of apiosylated sugar chains and glycosides. Here, we established a method for stabilizing UDP-apiose using bulky cations as counter ions. Bulky cations such as triethylamine effectively suppressed the degradation of UDP-apiose in solution. The half-life of UDP-apiose was increased to 48.1 ± 2.4 h at pH 6.0 and 25 °C using triethylamine as a counter cation. UDP-apiose coordinated with a counter cation enabled long-term storage under freezing conditions. UDP-apiose was utilized as a donor substrate for apigenin 7-O-β-D-glucoside apiosyltransferase to produce the apiosylated glycoside apiin. This apiosyltransferase assay will be useful for identifying genes encoding apiosyltransferases.
UDP-apiose 是一种糖基供体底物，由于其分子内的自我环化能力而易发生变化，从而形成了阿洛呋喃糖基-1,2-环磷酸酯。因此，稳定 UDP-apiose 对于其可用性以及鉴定和表征参与阿洛糖基化糖链和糖苷生物合成的阿洛基转移酶是必不可少的。在这里，我们建立了一种使用大体积阳离子作为抗衡离子来稳定 UDP-apiose 的方法。大体积阳离子，如三乙胺，可有效抑制 UDP-apiose 在溶液中的降解。在 pH 值为 6.0 和 25°C 下，使用三乙胺作为抗衡阳离子，UDP-apiose 的半衰期延长至 48.1±2.4 h。与抗衡阳离子配位的 UDP-apiose 能够在冷冻条件下长期储存。UDP-apiose 被用作芹糖苷 7-O-β-D-葡萄糖基阿洛基转移酶的供体底物，以生成阿洛糖苷 apiin。这种阿洛基转移酶测定法将有助于鉴定编码阿洛基转移酶的基因。
