Murine factor H Co-produced in yeast with protein disulfide isomerase ameliorated C3 dysregulation in factor H-deficient mice
Author(s)
Type
Journal Article
Abstract
Recombinant human factor H (hFH) has potential for treating diseases linked to aberrant complement regulation including C3 glomerulopathy (C3G) and dry age-related macular degeneration. Murine FH (mFH), produced in the same host, is useful for pre-clinical investigations in mouse models of disease. An abundance of FH in plasma suggests high doses, and hence microbial production, will be needed. Previously, Pichia pastoris produced useful but modest quantities of hFH. Herein, a similar strategy yielded miniscule quantities of mFH. Since FH has 40 disulfide bonds, we created a P. pastoris strain containing a methanol-inducible codon-modified gene for protein-disulfide isomerase (PDI) and transformed this with codon-modified DNA encoding mFH under the same promoter. What had been barely detectable yields of mFH became multiple 10s of mg/L. Our PDI-overexpressing strain also boosted hFH overproduction, by about tenfold. These enhancements exceeded PDI-related production gains reported for other proteins, all of which contain fewer disulfide-stabilized domains. We optimized fermentation conditions, purified recombinant mFH, enzymatically trimmed down its (non-human) N-glycans, characterised its functions in vitro and administered it to mice. In FH-knockout mice, our de-glycosylated recombinant mFH had a shorter half-life and induced more anti-mFH antibodies than mouse serum-derived, natively glycosylated, mFH. Even sequential daily injections of recombinant mFH failed to restore wild-type levels of FH and C3 in mouse plasma beyond 24 hours after the first injection. Nevertheless, mFH functionality appeared to persist in the glomerular basement membrane because C3-fragment deposition here, a hallmark of C3G, remained significantly reduced throughout and beyond the ten-day dosing regimen.
Date Issued
2021-05-12
Date Acceptance
2021-04-22
Citation
Frontiers in Immunology, 2021, 12, pp.1-17
ISSN
1664-3224
Publisher
Frontiers Media
Start Page
1
End Page
17
Journal / Book Title
Frontiers in Immunology
Volume
12
Copyright Statement
© 2021 Kerr, Herbert, Makou, Abramczyk, Malik, Lomax-Browne, Yang, Pappworth, Denton, Richards, Marchbank, Pickering and Barlow. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
License URL
Sponsor
Wellcome Trust
Identifier
http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000654216800001&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=1ba7043ffcc86c417c072aa74d649202
Grant Number
212252/Z/18/Z
Subjects
Science & Technology
Life Sciences & Biomedicine
Immunology
complement system
factor H
C3 glomerulonephritis mouse model
therapeutic protein
C3 glomerulonephritis
chaperonin
Pichia pastoris
protein disulfide isomerase (PDI)
COMPLEMENT FACTOR-H
MACULAR DEGENERATION
ASSOCIATIONS
EXPRESSION
Publication Status
Published
Article Number
ARTN 681098
Date Publish Online
2021-05-12