Production of biologically active recombinant human lactoferrin in Aspergillus oryzae.
We report the production of recombinant human lactoferrin in Aspergillus oryzae. Expression of human lactoferrin (hLF), a 78 kD glycoprotein, was achieved by placing the cDNA under the control of the A. oryzae alpha-amylase promoter and the 3' flanking region of the A. niger glucoamylase gene. Using this system, hLF is expressed and secreted into the growth medium at levels up to 25 mg/l. The recombinant lactoferrin is indistinguishable from human milk lactoferrin with respect to its size, immunoreactivity, and iron-binding capacity. The recombinant protein appears to be appropriately N-linked glycosylated and correctly processed at the N-terminus by the A. oryzae secretory apparatus. Lactoferrin is the largest heterologous protein and the first mammalian glycoprotein expressed in the Aspergillus system to date. Hence, this expression system appears suitable for the large-scale production and secretion of biologically active mammalian glycoproteins.
Department of Cell Biology, Baylor College of Medicine, Houston, Texas 77030.
P P
PP
Ward
J Y
JY
Lo
M
M
Duke
G S
GS
May
D R
DR
Headon
O M
OM
Conneely
D000595:Amino Acid Sequence; D001236:Aspergillus oryzae / Q000235:genetics*; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular / Q000379:methods; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D007501:Iron / Q000378:metabolism; D007781:Lactoferrin / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D000516:alpha-Amylases / Q000235:genetics
D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D004247:DNA; D007501:Iron; D000516:alpha-Amylases; D007781:Lactoferrin