An intrinsically disordered radish vacuolar calcium-binding protein (RVCaB) showed cryoprotective activity for lactate dehydrogenase with its hydrophobic region.

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).
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