Expression and characterization of functional recombinant bovine follicle-stimulating hormone (boFSHalpha/beta) produced in the milk of transgenic rabbits.
Bovine follicle-stimulating hormone (boFSH) is a heterodimeric glycoprotein that belongs to the pituitary gonadotropins. Bioactive FSH is composed of alpha and beta subunits which require extensive N-glycosylation and sialylation. The mammary gland of transgenic livestock is an attractive source for the synthesis of post-translationally modified proteins. Two mammary gland-specific gene constructs with the cDNA for the boFSH alpha (boFSHalpha) and beta (boFSHbeta) subunits controlled by bovine alpha-s1 casein regulatory sequences were co-microinjected into fertilized rabbit oocytes. Two FSHalpha/FSHbeta double transgenic rabbit lines were established. The transgene expression was strictly lactation and mammary gland specific. Protein analysis revealed the presence of the boFSH heterodimer in the milk of transgenic rabbits showing a molecular weight similar to that of purified pituitary gland derived boFSH (boFSH-P). Subunit specific antibodies detected both polypeptides with the expected molecular sizes. Biochemical characterization demonstrated the expected isoelectric points of the recombinant boFSH. The presence of the post-translationally added terminal sialic acid residues was indicated by wheat germ agglutinin (WGA) lectin Western blotting. The biological activity of the recombinant mammary gland produced boFSH was determined using a FSH-dependent reporter cell line. The bioactivity of the recombinant boFSH was comparable to that of purified boFSH-P.
Coulibaly
S
S
Ludwig Boltzmann Institute for Immunogenetic, Cytogenetic and Molecular Genetic Research, Vienna, Austria.
Besenfelder
U
U
Miller
I
I
Zinovieva
N
N
Lassnig
C
C
Kotler
T
T
Jameson
J L
JL
Gemeiner
M
M
Müller
M
M
Brem
G
G
D000818:Animals; D030801:Animals, Genetically Modified; D002417:Cattle; D005640:Follicle Stimulating Hormone / Q000096:biosynthesis* / Q000235:genetics*; D020869:Gene Expression Profiling; D006863:Hydrogen-Ion Concentration; D008892:Milk / Q000378:metabolism*; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics*
D011994:Recombinant Proteins; D005640:Follicle Stimulating Hormone