Department of Food Science and Technology, Tokyo University of Marine Science and Technology, Minato-ku, Tokyo, 108-8477, Japan.
J Sci Food Agric. 2013 May;93(7):1737-42. doi: 10.1002/jsfa.5961. Epub 2012 Nov 23.
Sarcoplasmic calcium-binding proteins (SCPs) have recently been identified as crustacean allergens. However, information on their primary structures is very limited and no recombinant SCP (rSCP) as an alternative of natural SCP (nSCP) is available. This study was aimed to elucidate primary structures of SCPs from two species of Penaeus shrimp (black tiger shrimp and kuruma shrimp) by cDNA cloning and to produce a black tiger shrimp rSCP preparation that is comparable in IgE reactivity to nSCP.
The full-length cDNAs encoding black tiger shrimp and kuruma shrimp SCPs were successfully cloned. Both SCPs are composed of 193 amino acid residues and share more than 80% sequence identity with the known crustacean SCPs. The black tiger shrimp SCP was then expressed in Escherichia coli using the pFN6A (HQ) Flexi vector system. Enzyme-linked immunosorbent assay (ELISA) and inhibition ELISA experiments demonstrated that rSCP has the same IgE reactivity as nSCP.
Our results provide further evidence for the high sequence identity among crustacean SCPs. In addition, rSCP will be a useful tool in studying crustacean allergens and also in the diagnosis of crustacean allergy.
肌浆钙结合蛋白（SCPs）最近被鉴定为甲壳类过敏原。然而，关于它们的一级结构的信息非常有限，并且没有重组 SCP（rSCP）作为天然 SCP（nSCP）的替代品。本研究旨在通过 cDNA 克隆阐明两种对虾（黑虎虾和斑节对虾）的 SCP 的一级结构，并制备与 nSCP 具有可比 IgE 反应性的黑虎虾 rSCP 制剂。
成功克隆了编码黑虎虾和斑节对虾 SCP 的全长 cDNA。两种 SCP 均由 193 个氨基酸残基组成，与已知的甲壳类 SCP 具有超过 80%的序列同一性。然后，使用 pFN6A（HQ）Flexi 载体系统在大肠杆菌中表达黑虎虾 SCP。酶联免疫吸附试验（ELISA）和抑制 ELISA 实验表明，rSCP 与 nSCP 具有相同的 IgE 反应性。
我们的结果进一步证明了甲壳类 SCP 之间具有很高的序列同一性。此外，rSCP 将成为研究甲壳类过敏原和甲壳类过敏诊断的有用工具。
