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- Title
Isolation of a cDNA Encoding a Protease from Perinereis aibuhitensis Grube.
- Authors
Rong-Gui LI; Dong-Meng QIAN; Dao-Sen GUO; Gui-Cai DU; Zhi-Yong YAN; Bin WANG
- Abstract
The cDNA encoding a protease of Perinereis aibuhitensis Grube (PPA) was cloned. The deduced amino acid sequence analysis showed that the protein had 49% identity to the C-terminal amino acid 169–246 of serine protease of Heterodera glycines. Northern blotting analysis indicated that the cDNA could hybridize with mRNA of approximately 260 bases isolated from the marine earthworm. The cDNA was amplified by polymerase chain reaction and cloned into pMAL-p2 to construct expression vector pMAL-PPA. pMAL-PPA was introduced into Escherichia coli BL21(DE3) and overexpression of PPA fused with maltose binding protein was achieved by isopropyl-β- D-thiogalactopyranoside induction. The fusion protein was purified by affinity chromatography on an amylose resin column and ion-exchange chromatography on a diethylaminoethyl-Sepharose 4B column. Rabbits were immunized with the purified protein and antiserum was prepared. The antibody could react with a protein of approximately 9 kDa extracted from the marine earthworm as shown by Western blotting analysis. The activity analysis of the recombinant PPA suggested that it was probably a plasminogen activator. Edited by Hernardez-Crespo PEDRO
- Subjects
PROTEOLYTIC enzymes; AMINO acids; PROTEINS; SOYBEAN cyst nematode; MESSENGER RNA; EARTHWORMS; BACTERIAL genetics; ESCHERICHIA coli
- Publication
Acta Biochimica et Biophysica Sinica, 2006, Vol 38, Issue 8, p543
- ISSN
1672-9145
- Publication type
Article
- DOI
10.1111/j.1745-7270.2006.00192.x