Efficient antibody production upon suppression of O mannosylation in the yeast Ogataea minuta.
When antibodies were expressed in the methylotrophic yeast Ogataea minuta, we found that abnormal O mannosylation occurred in the secreted antibody. Yeast-specific O mannosylation is initiated by the addition of mannose at serine (Ser) or threonine (Thr) residues in the endoplasmic reticulum via protein O mannosyltransferase (Pmt) activity. To suppress the addition of O-linked sugar chains on antibodies, we examined the possibility of inhibiting Pmt activity by the addition of a Pmt inhibitor during cultivation. The Pmt inhibitor was found to partially suppress the O mannosylation on the antibodies. Surprisingly, the suppression of O mannosylation was associated with an increased amount of assembled antibody (H2L2) and enhanced the antigen-binding activity of the secreted antibody. In this study, we demonstrated the expression of human antibody in O. minuta and elucidated the relationship between O mannosylation and antibody production in yeast.
Kuroda
Kousuke
K
National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba Central 6, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, Japan.
Kobayashi
Kazuo
K
Kitagawa
Yoshinori
Y
Nakagawa
Taishiro
T
Tsumura
Haruhiko
H
Komeda
Toshihiro
T
Shinmi
Daisuke
D
Mori
Eiji
E
Motoki
Kazuhiro
K
Fuju
Kazumi
K
Sakai
Teruyuki
T
Nonaka
Koichi
K
Suzuki
Takeshi
T
Ichikawa
Kimihisa
K
Chiba
Yasunori
Y
Jigami
Yoshifumi
Y
D000906:Antibodies / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D006031:Glycosylation / Q000187:drug effects; D006801:Humans; D008358:Mannose / Q000378:metabolism*; D008364:Mannosyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008954:Models, Biological; D011994:Recombinant Proteins / Q000096:biosynthesis; D015003:Yeasts / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*
D000906:Antibodies; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D008364:Mannosyltransferases; C082696:protein O-mannosyltransferase; D008358:Mannose