Molecular cloning and expression analysis of chymotrypsin-like serine protease from the Chinese shrimp, Fenneropenaeus chinensis.
A new member of the serine protease (SP) chymotrypsin (designated Fc-chy) was isolated from the hepatopancreas of Chinese shrimp Fenneropenaeus chinensis. The full-length cDNA of Fc-chy contained 951 nucleotides with a polyadenylation sequence and a polyA tail. It encoded a peptide of 271 amino acids with a signal peptide of 17 amino acids and an activation peptide of 28 amino acids. The mature peptide concludes 226 amino acids. It contained the conserved catalytic triad (H, D, and S). Similarity analysis showed that Fc-chy shared high identity with chymotrypsins from the Pacific white shrimp, Litopenaeus vannamei. Northern blot, quantitative real-time PCR, in situ hybridization, and western blot analysis were carried out to analyze the expression pattern and distribution profiles of Fc-chy after bacteria and virus challenges. The results showed that Fc-chy transcription and Fc-chy protein levels were upregulated in the hepatopancreas after bacterial and viral infection. Fc-chy from the hepatopancreas was purified by affinity chromatography. It showed high hydrolytic activity toward the substrate N-succinyl-ala-ala-pro-phe p-nitroanilide (AAPF), and its activity was inhibited by Kazal-type SP inhibitor from Chinese shrimp. All of these may indicate that Fc-chy is involved in the innate immune reactions in Chinese shrimp.
Shi
Xiu-Zhen
XZ
School of Life Sciences, Shandong University, Jinan, Shandong 250100, China.
Zhao
Xiao-Fan
XF
Wang
Jin-Xing
JX
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D005786:Gene Expression Regulation / Q000502:physiology*; D008969:Molecular Sequence Data; D033561:Penaeidae / Q000201:enzymology*; D010802:Phylogeny; D012697:Serine Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D012697:Serine Endopeptidases