The mitochondrial manganese superoxide dismutase gene in Chinese shrimp Fenneropenaeus chinensis: cloning, distribution and expression.
Manganese superoxide dismutase (MnSOD) plays an important role in crustacean immune defense reaction by eliminating oxidative stress. Knowledge on MnSOD at molecular level allows us to understand its regulatory mechanism in crustacean immune system. A novel mitochondrial manganese superoxide dismutase (mMnSOD) was cloned from hepatopancreas of Chinese shrimp Fenneropenaeus chinensis by 3' and 5' rapid amplification of cDNA ends (RACE) PCR. The full-length cDNA consists of 1185bp with a 660bp open reading frame, encoding 220 amino acids. The deduced amino acid sequence contains a putative signal peptide of 20 amino acids. Sequence comparison showed that the mMnSOD of F. chinensis shares 88% and 82% identity with that of giant freshwater prawn Macrobrachium rosenbergii and blue crab Callinectes sapidus, respectively. mMnSOD transcripts were detected in hepatopancreas, hemocytes, lymphoid organ, intestine, ovary, muscle and gill by Northern blotting. RT-PCR analysis indicated that mMnSOD showed different expression profiles in shrimp hemocytes and hepatopancreas after artificial infection with white spot syndrome virus (WSSV). In addition, a fusion protein containing mMnSOD was produced in vitro. LC-ESI-MS analysis showed that two peptide fragments (-GDVNTVISLAPALK- and -NVRPDYVNAIWK-) of the recombinant protein were identical to the corresponding sequence of M. rosenbergii mMnSOD, and the enzyme activity of the refolded recombinant protein was also measured.
Zhang
Qingli
Q
Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao, Shandong 266071, PR China.
Li
Fuhua
F
Wang
Bing
B
Zhang
Jiquan
J
Liu
Yichen
Y
Zhou
Qian
Q
Xiang
Jianhai
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D020869:Gene Expression Profiling; D043143:Hepatopancreas / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D033561:Penaeidae / Q000201:enzymology / Q000235:genetics*; D010802:Phylogeny; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D021241:Spectrometry, Mass, Electrospray Ionization; D013482:Superoxide Dismutase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D024101:Mitochondrial Proteins; D011993:Recombinant Fusion Proteins; D013482:Superoxide Dismutase