Human intrinsic factor expressed in the plant Arabidopsis thaliana.
Intrinsic factor (IF) is the gastric protein that promotes the intestinal uptake of vitamin B12. Gastric IF from animal sources is used in diagnostic tests and in vitamin pills. However, administration of animal IF to humans becomes disadvantageous because of possible pathogenic transmission and contamination by other B12 binders. We tested the use of recombinant plants for large-scale production of pathogen-free human recombinant IF. Human IF was successfully expressed in the recombinant plant Arabidopsis thaliana. Extract from fresh plants possessed high B12-binding capacity corresponding to 70 mg IF per 1 kg wet weight. The dried plants still retained 60% of the IF activity. The purified IF preparation consisted of a 50-kDa glycosylated protein with the N-terminal sequence of mature IF. Approximately one-third of the protein was cleaved at the internal site em leader PSNP downward arrow GPGP. The key properties of the preparation obtained were identical to those of native IF: the binding curves of vitamin B12 to recombinant IF and gastric IF were the same, as were those for a B12 analogue cobinamide, which binds to IF with low affinity. The absorbance spectra of the vitamin bound to recombinant IF and gastric IF were alike, as was the interaction of recombinant and native IF with the specific receptor cubilin. The data presented show that recombinant plants have a great potential as a large-scale source of human IF for analytical and therapeutic purposes.
Fedosov
Sergey N
SN
Protein Chemistry Laboratory, Department of Molecular and Structural Biology, University of Aarhus, Denmark.
Laursen
Niels B
NB
Nexø
Ebba
E
Moestrup
Søren K
SK
Petersen
Torben E
TE
Jensen
Erik Ø
EØ
Berglund
Lars
L
D016960:Agrobacterium tumefaciens; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D018014:Gene Transfer Techniques; D006801:Humans; D007437:Intrinsic Factor / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D030821:Plants, Genetically Modified; D011956:Receptors, Cell Surface / Q000378:metabolism; D013053:Spectrophotometry; D014805:Vitamin B 12 / Q000378:metabolism*
D011956:Receptors, Cell Surface; C029486:intrinsic factor-cobalamin receptor; D007437:Intrinsic Factor; D014805:Vitamin B 12