Olive flounder (Paralichthys olivaceus) cystatin C: cloning, mRNA expression, and enzymatic characterization of olive flounder cystatin C.
Appl Biochem Biotechnol
; 170(5): 1216-28, 2013 Jul.
Article
em En
| MEDLINE
| ID: mdl-23649306
ABSTRACT
Cystatins are endogenous inhibitors of mammalian lysosomal cysteine proteinases, such as cathepsins B, L, H, and S. Cystatin C belongs to the type 2 cystatin family. In this study, the 751-bp cystatin C cDNA (PoCystatin C) of olive flounder (Paralichthys olivaceus) was cloned by screening from the olive flounder cDNA library. The mRNA expression of the PoCystatin C gene was examined in various tissues from normal and lipopolysaccharide (LPS)-stimulated olive flounder by RT-PCR and was compared with inflammatory cytokines IL-1ß, IL-6, and IL-8. PoCystatin C transcripts ubiquitously existed in all normal and LPS-stimulated tissues that were tested. The recombinant PoCystatin C protein was expressed in Escherichia coli BL21(DE3) in pCold™ TF DNA expression vector as a 70-kDa fusion protein. The protease inhibitory activities of recombinant PoCystatin C toward papain cysteine protease, piscine cathepsins (L, S, K, F, and X), and bovine cathepsin B were measured with the synthetic fluorogenic peptide substrates. PoCystatin C tightly inhibited papain cysteine protease, whereas cathepsins L, S, K, F, X, and B were inhibited with lower affinities. Our results indicate that the P. olivaceus cystatin C is a homolog of mammalian cystatin C due to its sequence, structure, tissue expression, and biochemical activity.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Peptídeo Hidrolases
/
Inibidores de Proteases
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Linguado
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RNA Mensageiro
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Cistatina C
Limite:
Animals
Idioma:
En
Revista:
Appl Biochem Biotechnol
Ano de publicação:
2013
Tipo de documento:
Article
País de afiliação:
Estados Unidos