Expression in Escherchia coli and purification of sea bass (Dicentrarchus labrax) interleukin 1beta, a possible immunoadjuvant in aquaculture.
Interleukin 1beta (IL-1beta) is a pleiotropic cytokine that plays a pivotal role in regulating immune responses. Our group has recently cloned IL-1beta from sea bass (Dicentrarchus labrax), one of the main Mediterranean aquacultured fish species. The cDNA is 1292 bp and codes for a deduced peptide of 29.4 kDa with a pI of 5.1. As for trout and carp IL-1beta precursor sequence, no candidate cut site for ICE (IL-1beta converting enzyme) enzyme was apparent in the alignments of sea bass IL-1beta with other mammalian IL-1betas. Nevertheless, a possible mature peptide could start at Ala86, giving a protein of 176 amino acids. The nucleotide sequence coding for this polypeptide was cloned into a pQE-30 expression vector. The plasmid was then transformed in Escherichia coli, and the recombinant protein was purified. Finally, we demonstrated that this purified recombinant IL-1beta was able to induce IL-1beta gene expression in a dose-dependent manner on cells purified from sea bass head kidney and could have immunoadjuvant effects in sea bass vaccination experiments.
Buonocore
Francesco
F
Dipartimento di Scienze Ambientali, Università della Tuscia, 01100 Viterbo, Italy. fbuono@unitus.it
Mazzini
Massimo
M
Forlenza
Maria
M
Randelli
Elisa
E
Secombes
Christopher J
CJ
Zou
Jun
J
Scapigliati
Giuseppe
G
D000818:Animals; D017756:Aquaculture; D001492:Bass / Q000235:genetics* / Q000276:immunology; D015153:Blotting, Western; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000378:metabolism; D005786:Gene Expression Regulation; D007136:Immunoglobulins / Q000097:blood; D007375:Interleukin-1 / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014733:Vibrio / Q000276:immunology
D017931:DNA Primers; D018076:DNA, Complementary; D007136:Immunoglobulins; D007375:Interleukin-1