Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea.
Department of Marine Life Sciences & Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province, 63243, Republic of Korea.
Fish Shellfish Immunol. 2019 May;88:364-374. doi: 10.1016/j.fsi.2019.03.010. Epub 2019 Mar 7.
The fourth member of the typical 2-cysteine peroxiredoxin (Prx4) is a well-known antioxidant enzyme, which reduces different peroxides in their catalytic process. The present study reports the identification of the rockfish Sebastes schlegelii Prx4 (SsPrx4) at a genomic level, as well as the characterization of its structural and functional features. SsPrx4 harbors a complete ORF of 786 bp encoding a polypeptide (29 kDa) of 262 amino acids (aa) with an isoelectric point of 6.2. Thioredoxin 2 domain was prominent in the SsPrx4 sequence, which has a signal peptide (31 bp) at the N-terminus. Hence, the SsPrx4 may be functionally active in the cytoplasm of rockfish cells. Moreover, two VCP motifs and three catalytic triad residues (T, C, R) were identified in the SsPrx4 protein sequence. A peroxidatic cysteine (C) and resolving cysteines (C) were detected at the VCP motifs. The rockfish Prx4 genome consists of seven exons, which are similar to the architecture of other Prx4 orthologs. The deduced amino acid sequence of SsPrx4 shares a relatively high amino acid sequence identity (91.6%) and close evolutionary relationship with Miichthys miiuy and Stegastes partitus Prx4. The potential for scavenging extracellular HO was evidenced by the purified recombinant SsPrx4 protein (rSsPrx4) in vitro system. Moreover, rSsPrx4 may protect the plasmid DNA in a metal-catalyzed oxidation system and catalyze the reduction of an insulin disulfide bond. Quantitative real-time PCR revealed that SsPrx4 mRNA was ubiquitously expressed in fourteen different tissues, with the highest expression observed in the liver followed by the ovary, and kidney tissues. Transcriptional modulations were observed in liver and spleen tissues of rockfish after injecting them with bacterial stimuli, including Streptococcus iniae, LPS, and a viral mimic of poly I:C. Together, the results suggest that SsPrx4 may play an important role in both the antioxidant and innate immune defense of black rockfish. These findings provide structural and functional insights into the SsPrx4 of the teleost.
第四种典型的 2-半胱氨酸过氧化物酶（Prx4）是一种众所周知的抗氧化酶，它在催化过程中还原不同的过氧化物。本研究在基因组水平上报告了岩鱼Sebastes schlegelii Prx4（SsPrx4）的鉴定，以及其结构和功能特征的描述。SsPrx4 含有一个完整的 786bp 的 ORF，编码一个 29kDa 的多肽，由 262 个氨基酸（aa）组成，等电点为 6.2。硫氧还蛋白 2 结构域在 SsPrx4 序列中很突出，其 N 端有一个信号肽（31bp）。因此，SsPrx4 可能在岩鱼细胞的细胞质中具有功能活性。此外，在 SsPrx4 蛋白序列中鉴定出两个 VCP 基序和三个催化三联体残基（T、C、R）。在 VCP 基序中检测到过氧物酶半胱氨酸（C）和解析半胱氨酸（C）。岩鱼 Prx4 基因组由七个外显子组成，与其他 Prx4 同源物的结构相似。SsPrx4 的推导氨基酸序列与 Miichthys miiuy 和 Stegastes partitus Prx4 具有相对较高的氨基酸序列同一性（91.6%）和密切的进化关系。体外系统纯化的重组 SsPrx4 蛋白（rSsPrx4）证明了其清除细胞外 HO 的能力。此外，rSsPrx4 可能在金属催化氧化系统中保护质粒 DNA，并催化胰岛素二硫键的还原。定量实时 PCR 显示，SsPrx4 mRNA 在 14 种不同组织中广泛表达，肝脏中的表达最高，其次是卵巢和肾脏组织。在注射细菌刺激物（包括链球菌、LPS 和聚 I:C 的病毒模拟物）后，岩鱼的肝和脾组织中观察到转录调节。总之，这些结果表明 SsPrx4 可能在黑岩鱼的抗氧化和固有免疫防御中发挥重要作用。这些发现为硬骨鱼的 SsPrx4 提供了结构和功能上的见解。
