Purification, physicochemical and immunological characterization of arginine kinase, an allergen of crayfish (Procambarus clarkii)

Abstract

Arginine kinase (AK) has attracted considerable attention because it has been identified as a shellfish allergen. However, little information is available about AK in crayfish (Procambarus clarkii). In this study, crayfish AK was purified and cloned. Its physicochemical properties, processing stability, and immunological characteristics were analyzed. Crayfish AK was purified by column chromatography, which revealed a single band with molecular mass of 40kDa; this result was further confirmed by mass spectrometry. The full-length gene sequence of crayfish AK was 1462bp and encoded a protein of 357 amino acid residues. The results of this study revealed that crayfish AK is a glycoprotein with an isoelectric point of approximately 6.5. Thermal stability assays revealed that crayfish AK easily forms aggregates at temperatures >44°C and was stable at pH 4.0–8.0. SDS–PAGE and dot blotting were used to assess processing stability of purified AK. The results revealed that the IgE-binding activity of crayfish AK is reduced after boiling.