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Production and purification of biologically active recombinant tilapia (Oreochromis niloticus) prolactins
Swennen, D.; Rentier-Delrue, Françoise; Auperin, B. et al.
1991In Journal of Endocrinology, 131 (2), p. 219-27
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Keywords :
Animals; Blotting, Western; Electrophoresis, Polyacrylamide Gel; Fishes/*metabolism; Kidney/metabolism; Plasmids; Prolactin/*biosynthesis/isolation & purification/metabolism; Protein Binding; Protein Engineering/methods; Recombinant Proteins/*biosynthesis/isolation & purification/metabolism
Abstract :
[en] Recombinant expression vectors carrying tilapia prolactin-I or -II (tiPRL-I or tiPRL-II) cDNA were constructed and the tiPRL-I and II proteins were produced in E. coli as inclusion bodies. These inclusion bodies were dissolved in 6 mol urea/l. Refolding of the proteins was followed by SDS-PAGE under non-reducing conditions so as to visualize the oxidized state of the molecules. Proteins tiPRL-I and tiPRL-II were purified by gel filtration and ion-exchange chromatography. The N-terminal sequence and bioactivities of both purified proteins were then analysed. Recombinant tiPRL-I and tiPRL-II induced a significant rise in plasma calcium levels as well as in mucocyte density in the abdominal skin epithelium. When tested on kidney membrane, both proteins exhibited potency in competing with 125I-labelled tiPRL-I for binding sites, but tiPRL-I seemed to be more potent than tiPRL-II in competing for these sites. The results obtained for the biological activities tested suggest that both recombinant prolactins were correctly refolded and had retained the full biological activity previously observed with the natural hormone preparations extracted from the animals.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Swennen, D.
Rentier-Delrue, Françoise ;  Université de Liège - ULiège > Département des sciences de la vie > Biologie et génétique moléculaire - GIGA-R : Coordination scientifique
Auperin, B.
Prunet, P.
Flik, G.
Wendelaar Bonga, S. E.
Lion, Michelle ;  Université de Liège - ULiège > Département des sciences de la vie > GIGA-R : Biologie et génétique moléculaire
Martial, Joseph ;  Université de Liège - ULiège > Département des sciences de la vie > GIGA-R : Biologie et génétique moléculaire
Language :
English
Title :
Production and purification of biologically active recombinant tilapia (Oreochromis niloticus) prolactins
Publication date :
1991
Journal title :
Journal of Endocrinology
ISSN :
0022-0795
eISSN :
1479-6805
Publisher :
Society for Endocrinology, Bristol, United Kingdom
Volume :
131
Issue :
2
Pages :
219-27
Peer reviewed :
Peer Reviewed verified by ORBi
Available on ORBi :
since 27 August 2009

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