Modified host cells and hybrid oligosaccharides for use in bioconjugate production
Provided herein are host cells capable of producing hybrid oligosaccharides and polysaccharides, wherein said hybrid oligosaccharides and polysaccharides do not comprise a hexose at the reducing end of their first repeat unit. Also provided herein are hybrid oligosaccharides or polysaccharides and bioconjugates which can be produced by the host cells described herein, wherein said bioconjugates comprise a carrier protein linked to a hybrid oligosaccharide or polysaccharide that does not comprise a hexose at the reducing end of its first repeat unit.
1. A hybrid oligosaccharide or polysaccharide having a structure
(B) n -A→
a) wherein A is an oligosaccharide repeat unit (i) of a capsular saccharide of a Gram positive bacterial capsular saccharide, a Streptococcus pneumoniae capsular saccharide, (ii) that contains at least 2, 3, 4, 5, 6, 7 or 8 monosaccharides, (iii) with a hexose monosaccharide derivative at the reducing end (indicated by arrow);
b) wherein B is an oligosaccharide repeat unit (i) containing at least 2, 3, 4, 5, 6, 7 or 8 monosaccharides and (ii) with a hexose monosaccharide at the reducing end of the repeat optionally wherein the hexose monosaccharide comprises glucose, galactose, rhamnose, arabinotol, fucose, or mannose;
c) wherein A and B are different oligosaccharide repeat units; and
wherein n is at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20.
2. The hybrid oligosaccharide or polysaccharide of claim 1 , linked to a carrier protein.
3. The hybrid oligosaccharide or polysaccharide of claim 1 , wherein the hexose monosaccharide derivative is any monosaccharide in which C-2 position is modified with an acetamido group.
4. The hybrid oligosaccharide or polysaccharide of claim 3 , in which the hexose monosaccharide derivative is N-acetylglucosamine (GlcNAc), N-acetylgalactoseamine (GalNAc), 2,4-Diacetamido-2,4,6-trideoxyhexose (DATDH), N-acetylfucoseamine (FucNAc), or N-acetylquinovosamine (QuiNAc).
5. The hybrid oligosaccharide or polysaccharide of claim 1 , wherein (A) is an oligosaccharide repeat unit (i) of a capsular saccharide of a Gram positive bacterial capsular saccharide
Streptococcus pneumoniae capsular polysaccharide (CP)
CP15A with a hexose monosaccharide derivative at the reducing end (indicated by arrow).
6. The hybrid oligosaccharide or polysaccharide of claim 2 , wherein the carrier protein is detoxified Exotoxin A of P. aeruginosa (EPA), CRM197, maltose binding protein (MBP), Diphtheria toxoid, Tetanus toxoid, detoxified hemolysin A of S. aureus , clumping factor A, clumping factor B, E. coli FimH, E. coli FimHC, E. coli heat labile enterotoxin, detoxified variants of E. coli heat labile enterotoxin, Cholera toxin B subunit (CTB), cholera toxin, detoxified variants of cholera toxin, E. coli Sat protein, the passenger domain of E. coli Sat protein, Streptococcus pneumoniae Pneumolysin and detoxified variants thereof, C. jejuni AcrA, a C. jejuni natural glycoprotein, PcrV (aka LcrV, EspA, SseB), PopB (YopB, YopD, FliC), or OprF, Oprl.
7. A bioconjugate comprising the hybrid oligosaccharide or polysaccharide of claim 1 and a carrier protein N-linked to an oligosaccharide or polysaccharide, produced by a method comprising (i) culturing a host cell expressing said bioconjugates under conditions suitable for the production of proteins and (ii) isolating said bioconjugate.
8. A composition comprising the hybrid oligosaccharide or polysaccharide of claim 2 .
9. A method of treating Streptococcus pneumoniae in a subject, comprising administering to a subject the composition of claim 8 .
10. A method of inducing an immune response against Streptococcus pneumoniae in a subject, comprising administering to a subject the composition of claim 8 .