Molecular cloning and functional expression of hemolysin from the sea anemone Actineria villosa.
Protein Expr Purif
; 40(2): 379-84, 2005 Apr.
Article
em En
| MEDLINE
| ID: mdl-15766880
ABSTRACT
The full-length cDNA that encodes the hemolytic toxin Avt-I, with 226 amino acids, from the venomous sea anemone Actineria villosa has been cloned using the oligo-capping method. The cDNA contains 681bp open reading frame and its predicted amino acid sequences revealed that Avt-I was basic polypeptides without cysteine residues and Arg-Gly-Asp (RGD) motif sequence. The mature Avt-I has a predicted molecular weight of 19.6 kDa and its theoretical isoelectric point is 9.3. The Avt-I revealed 99, 61, 57, and 57% amino acid similarity with hemolytic toxins Pstx20, EqtII, StII, and HmT from Phyllodiscus semoni, Actinia Equina, Stichodactyla helianthus, and Heteractis magnifica, respectively. The characteristic amphiphilic alpha-helix structure was found at the N-terminal region of the mature Avt-I. Recombinant Avt-I (rAvt-I) was expressed in Escherichia coli BL21 (DE3) strain as a biologically active form and purified rAvt-I caused 50% hemolytic activity against 1% sheep erythrocytes at a concentration of 6.3 ng/ml (0.32 nM). M9Y medium led to more than 2-fold increase in rAvt-I yield than cultivation in Luria-Bertani medium.
Buscar no Google
Base de dados:
MEDLINE
Assunto principal:
Anêmonas-do-Mar
/
Clonagem Molecular
/
Proteínas Hemolisinas
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Revista:
Protein Expr Purif
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2005
Tipo de documento:
Article
País de afiliação:
Japão