Production of biologically active salmon calcitonin in the milk of transgenic rabbits.
Salmon calcitonin (sCT) is an example of one of the many bioactive peptides that require amidation of the carboxy terminus for full potency. We describe a method for the production of amidated sCT in the mammary gland of transgenic rabbits. Expression of a fusion protein comprising human alpha lactalbumin joined by an enterokinase cleavable linker to sCT was directed to the mammary gland under the control of the ovine beta lactoglobulin promoter. C-terminal amidation in vivo was achieved by extending the sCT by a single glycine residue that provides a substrate for endogenous amidating activity in the mammary gland. Full characterization of the released sCT demonstrated it to be equivalent to synthetic standard in terms of structure, purity, and potency.
McKee
C
C
PPL Therapeutics Ltd., Roslin, Edinburgh, Scotland. McKee@pplros.demon.co.uk
Gibson
A
A
Dalrymple
M
M
Emslie
L
L
Garner
I
I
Cottingham
I
I
D000577:Amides / Q000737:chemistry; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D002116:Calcitonin / Q000096:biosynthesis* / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid; D004247:DNA / Q000737:chemistry; D005260:Female; D007768:Lactalbumin / Q000737:chemistry; D008321:Mammary Glands, Animal / Q000378:metabolism; D013058:Mass Spectrometry; D008892:Milk / Q000737:chemistry*; D008969:Molecular Sequence Data; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000737:chemistry
D000577:Amides; D011993:Recombinant Fusion Proteins; C028815:salmon calcitonin; D002116:Calcitonin; D004247:DNA; D007768:Lactalbumin