Graduate School of Integrated Science and Technology, Shizuoka University, 836 Ohya, Shizuoka, Shizuoka 422-8529, Japan.
Research Institute of Green Science and Technology, Shizuoka University, 836 Ohya, Shizuoka, Shizuoka 422-8529, Japan; Graduate School of Integrated Science and Technology, Shizuoka University, 836 Ohya, Shizuoka, Shizuoka 422-8529, Japan; Graduate School of Science and Technology, Shizuoka University, 836 Ohya, Shizuoka, Shizuoka 422-8529, Japan.
Int J Biol Macromol. 2021 Jul 1;182:1130-1137. doi: 10.1016/j.ijbiomac.2021.04.056. Epub 2021 Apr 20.
A soluble protein fraction from radish (Raphanus sativus L.) taproot had cryoprotective activity for lactate dehydrogenase (LDH). The activity was found mainly in the heat-stable fractions of soluble proteins. The cryoprotective protein, whose molecular mass was 43 kDa in sodium dodecyl sulfate polyacrylamide gel electrophoresis, was obtained by successive chromatographies on TOYOPEARL SuperQ and TOYOPEARL DEAE. MALDI-TOF MS/MS analysis indicated that the purified protein was a radish vacuolar calcium-binding protein (RVCaB), which is reportedly related to calcium storage in the vacuoles of radish taproot. The purified RVCaB inhibited the cryoinactivation, cryodenaturation, and cryoaggregation of LDH. RVCaB had greater cryoprotective activity than general cryoprotectants. When RVCaB was divided into 15 segments (Seg01 to Seg15, 15 amino acids each), Seg03, which had a high hydrophobicity scale, showed remarkable cryoprotective activity. This indicated that RVCaB protected LDH from freezing and thawing damage presumably through a specific hydrophobic area (i.e., Seg03).
萝卜（Raphanus sativus L.）主根的一种可溶性蛋白质部分具有乳酸脱氢酶（LDH）的冷冻保护活性。该活性主要存在于可溶性蛋白质的热稳定部分中。该冷冻保护蛋白在十二烷基硫酸钠聚丙烯酰胺凝胶电泳中的分子量为 43 kDa，通过连续层析 TOYOPEARL SuperQ 和 TOYOPEARL DEAE 获得。MALDI-TOF MS/MS 分析表明，纯化的蛋白质是一种萝卜液泡钙结合蛋白（RVCaB），据报道与萝卜主根液泡中的钙储存有关。纯化的 RVCaB 抑制了 LDH 的冷冻失活、冷冻变性和冷冻聚集。RVCaB 比一般的冷冻保护剂具有更强的冷冻保护活性。当 RVCaB 被分成 15 个片段（Seg01 到 Seg15，每个片段 15 个氨基酸）时，具有高疏水性尺度的 Seg03 显示出显著的冷冻保护活性。这表明 RVCaB 可能通过特定的疏水区（即 Seg03）保护 LDH 免受冷冻和解冻损伤。
