Fucosidase from bacteroides and methods using the same
The present disclosure relates to an α-fucosidase having α-(1,2), α-(1,3), α-(1,4), and α-(1,6) fucosidase activity. The present disclosure also relates to the compositions comprising the α-fucosidase, and the methods of producing and using the α-fucosidase in cleaving α-(1,2), α-(1,3), α-(1,4), and/or α-(1,6)-linked fucoses in the glycoconjugates.
1. A composition comprising an α-fucosidase, wherein the composition comprises:
(a) an α-fucosidase polypeptide having the sequence of SEQ ID NO: 1;
(b) at least one glycosidase selected from the group consisting of Endo-beta-N-acetylglucosaminidases (NAG), EndoA, EndoF1, EndoF2, EndoF3, EndoH, EndoM, EndoS, and variants thereof; and
(c) a glycoprotein comprising an N-linked glycan, wherein the glycoprotein is selected from the group consisting of an immunoglobulin G, an Fc-fusion protein, erythropoietin, interferon and influenza hemagglutinin.
2. The composition of claim 1 , wherein the α-fucosidase is an isolated polypeptide.
3. The composition of claim 1 , wherein the α-fucosidase is a recombinant Bacteroides α-L-fucosidase.
4. The composition of claim 1 , wherein the α-fucosidase can hydrolyze α-(1,2), α-(1,3), α-(1,4), and α-(1,6)-linked fucoses present in N- and/or O-linked glycans in a glycoconjugate.
5. The composition of claim 1 , wherein the α-fucosidase has a pH optimum from about pH 7.0 to about pH 7.5.
6. The composition of claim 1 , wherein the immunoglobulin G comprises an anti-CD20 antibody or an anti-TNFα antibody.
7. The composition of claim 6 , wherein the anti-CD20 antibody is a chimeric IgG1.
8. The composition of claim 7 , wherein the anti-CD20 antibody is rituximab.
9. The composition of claim 6 , wherein the anti-TNFα antibody is adalimumab.
10. A composition comprising an α-fucosidase wherein the composition comprises:
(a) an α-fucosidase polypeptide having the amino acid sequence of SEQ ID NO: 1;
(b) at least one glycosidase selected from the group consisting of Endo-beta-N-acetylglucosaminidases (NAG), EndoA, EndoF1, EndoF2, EndoF3, EndoH, EndoM, EndoS, and variants thereof; and
(c) a glycoprotein selected from the group consisting of an immunoglobulin G, a Fc-fusion protein, erythropoietin, interferon and influenza hemagglutinin.
11. The composition of claim 10 , wherein α-fucosidase is a recombinant Bacteroides α-L-fucosidase.
12. The composition of claim 10 , wherein the α-fucosidase can hydrolyze α-(1,2), α-(1,3), α-(1,4), and α-(1,6)-linked fucoses present in N- and/or O-linked glycans in a glycoconjugate.
13. A composition comprising an α-fucosidase wherein the composition comprises:
(a) an α-fucosidase polypeptide having the sequence of SEQ ID NO: 1;
(b) at least one glycosidase selected from the group consisting of Endo-beta-N-acetylglucosaminidases (NAG), EndoA, EndoF1, EndoF2, EndoF3, EndoH, EndoM, EndoS, and variants thereof; and
(c) a glycoprotein comprising an N-linked glycan, wherein the glycoprotein is selected from the group consisting of an immunoglobulin G, an Fc-fusion protein, erythropoietin, interferon and influenza hemagglutinin; and wherein the pH of the composition is between about 7.0 to about 8.0.
14. A composition comprising an α-fucosidase wherein the composition comprises:
(a) an α-fucosidase polypeptide having the sequence of SEQ ID NO: 1;
(b) at least one glycosidase selected from the group consisting of Endo-beta-N-acetylglucosaminidases (NAG), EndoA, EndoF1, EndoF2, EndoF3, EndoH, EndoM, EndoS, and variants thereof; and
(c) a glycoprotein comprising an N-linked glycan, wherein the glycoprotein is selected from the group consisting of an immunoglobulin G, an Fc-fusion protein, erythropoietin, interferon and influenza hemagglutinin; and wherein the composition excludes a divalent cation selected from the group consisting of Zn 2+ , Cu 2+ and Ni 2+ .
15. A composition comprising an α-fucosidase, wherein the composition comprises:
(a) an α-fucosidase polypeptide having the sequence of SEQ ID NO: 1;
(b) at least one glycosidase selected from the group consisting of EndoF1, EndoF2, EndoF3, EndoH, EndoS, and variants thereof; and
(c) a glycoprotein comprising an N-linked glycan, wherein the glycoprotein is selected from the group consisting of an immunoglobulin G, an Fc-fusion protein, erythropoietin, interferon and influenza hemagglutinin.
16. A composition comprising an α-fucosidase, wherein the composition comprises:
(a) an α-fucosidase polypeptide having the sequence of SEQ ID NO: 1; and
(b) (i) Endo S and optionally at least one glycosidase selected from the group consisting of EndoF1, EndoF2, EndoF3, EndoH, EndoS, and variants thereof; or (ii) a glycosidase cocktail comprising Endo F1, Endo F2, Endo F3, and Endo H; and
(c) an immunoglobulin G or an Fc-fusion protein comprising an N-linked glycan.
17. A composition made by a process comprising:
contacting a glycoprotein with an α-fucosidase to remove one or more fucoses in the glycoprotein,
wherein the composition comprises an α-fucosidase polypeptide having the sequence of SEQ ID NO: 1 and a glycosidase cocktail comprising EndoF1, EndoF2, EndoF3, EndoH and EndoM,
wherein the glycoprotein comprises an N-linked glycan and is selected from the group consisting of an immunoglobulin G, an Fc-fusion protein, erythropoietin, interferon and influenza hemagglutinin.