Process for the modification of a glycoprotein using a β-(1,4)-N-acetylgalactosaminyl transferase or a mutant thereof

1. A process for producing a modified glycoprotein, comprising contacting a glycoprotein comprising a glycan comprising a terminal GlcNAc moiety, with a sugar-derivative nucleotide Su(A)-Nuc, in the presence of a β-(1,4)-N-acetylgalactosaminyltransferase having at least 90% identity to the sequence selected from the group consisting of SEQ ID NO:2, 3, 4, 5, 6, 7, 8, 9, 10, and 14, wherein:

the glycan comprising a terminal GlcNAc-moiety is of formula (1) or (2):

wherein:

GlcNAc is N-acetyl-glucosaminyl;

Fuc is fucosyl;

b is 0 or 1;

d is 0 or 1;

e is 0 or 1; and

G is a monosaccharide, or a linear or branched oligosaccharide comprising 2 to 20 sugar moieties; and

the sugar-derivative nucleotide Su(A)-Nuc is of formula (3):

wherein:

a is 0 or 1;

Nuc is a nucleotide;

U is [C(R 1 ) 2 ] n or [C(R 1 ) 2 ] p —O—[C(R 1 ) 2 C(R 1 ) 2 O] o —[C(R 1 ) 2 ] q , wherein n is an integer in the range of 0 to 24; o is an integer in the range of 0 to 12; q and p are independently 0, 1 or 2; and R 1 is independently selected from the group consisting of H, F, Cl, Br, I and an optionally substituted C 1 -C 24 alkyl group;

T is a C 3 -C 12 (hetero)arylene group, wherein the (hetero)arylene group is optionally substituted; and

A is selected from the group consisting of:

(a) —N 3 ;

(b) —C(O)R 3 , wherein R 3 is an optionally substituted C 1 -C 24 alkyl group;

(c) C≡C—R 4 , wherein R 4 is hydrogen or an optionally substituted C 1 -C 24 alkyl group;

(d) —SH;

(e) SC(O)R 8 , wherein R 8 is an optionally substituted C 1 -C 24 alkyl group;

(f) —SC(V)OR 8 , wherein V is 0 or S, and R 8 is an optionally substituted C 1 -C 24 alkyl group;

(g) —X, wherein X is selected from the group consisting of F, Cl, Br and I;

(h) —OS(O) 2 R 5 , wherein R 5 is selected from the group consisting of C 1 -C 24 alkyl groups, C 6 -C 24 aryl groups, C 7 -C 24 alkylaryl groups and C 7 -C 24 arylalkyl groups, the alkyl groups, aryl groups, alkylaryl groups and arylalkyl groups being optionally substituted;

(i) R 11 , wherein R 11 is an optionally substituted C 2 -C 24 alkyl group;

(j) R 12 , wherein R 12 is an optionally substituted terminal C 2 -C 24 alkenyl group; and

(k) R 13 , wherein R 13 is an optionally substituted terminal C 3 -C 24 allenyl group.

2. The process of claim 1 , wherein when U is [C(R 1 ) 2 ] n , n is an integer in the range of 1 to 24, and when U is [C(R 1 ) 2 ] p —O—[C(R 1 ) 2 C(R 1 ) 2 O] o —[C(R 1 ) 2 ] q , o is an integer in the range of 1 to 12 and/or p is 1 or 2 and/or q is 1 or 2.

3. The process according to claim 1 , wherein the sugar-derivative nucleotide Su(A)-Nuc is of formula (9) or (10):

wherein Nuc, A, U and T are as defined in claim 1 .

4. The process of claim 1 , wherein the nucleotide is uridine-diphosphate (UDP).

5. The process of claim 1 , wherein the sugar-derivative nucleotide is of formula (17) or (18):

.

6. The process of claim 1 , wherein the glycan comprising a terminal GlcNAc moiety is of formula (1), (26) or (27):

wherein:

b is as defined in claim 1 .

7. The process of claim 6 , wherein the glycoprotein comprising a glycan comprising a terminal GlcNAc moiety is an antibody.

8. The process of claim 6 , wherein the β-(1,4)-N-acetylgalactosaminyltransferase is or is derived from an invertebrate β-(1,4)-GalNAcT enzyme.

9. The process of claim 8 , wherein the invertebrate is selected from the group consisting of Caenorhabditis elegans, Caenorhabditis remanei, Caenorhabditis briggsae, Ascaris suum, Trichoplusia ni, Drosophila melanogaster, Wuchereria bancrofti, Loa loa, Cerapachys biroi, Zootermopsis nevadensis, Camponotus floridanus, Crassostrea gigas , and Danaus plexippus.

10. The process of claim 9 , wherein the invertebrate is selected from the group consisting of Caenorhabditis elegans, Ascaris suum, Trichoplusia ni , and Drosophila melanogaster.

11. The process of claim 6 , wherein A is —N 3 .