| cysteine conjugation |
fast conjugation
reactions; minimal structural perturbation |
requires prereduction
and may require reoxidation |
global reduction/alkylation |
DAR of 4 or 8 depending on linker chemistry |
| cysteine to serine mutation |
DAR of 2, 4, or 6 |
| THIOMAB |
maleimide conjugates unstable,
but newer chemistries exist |
| N-terminal cysteine conjugation |
oxazolidine conjugates intentionally
unstable |
| |
|
|
|
|
| glycoconjugation |
no protein
engineering required |
site of modification
is immutable; DARs tend to be lower due to glycan heterogeneity |
periodate oxidation of fucose
or sialic acid |
methionine
oxidation may
be problematic |
| enzymatic transfer of azidosugars |
asymmetric cyclooctynes
yield two regioisomers |
| metabolic incorporation
of thiofucose |
DAR
limited by metabolic
incorporation efficiency |
| |
|
|
|
|
| unnatural or
noncanonical amino acid incorporation |
minimal structural
perturbation; potentially enables wide variety of bioorthogonal ligation
reactions |
technically
complicated |
amber
codon suppression |
aryl oxime ligation is slow |
| cell-free amber codon suppression |
can quickly screen variants;
generates aglycosylated antibody |
| selenocysteine incorporation |
mostly conjugates with DAR of 1; C-terminal incorporation |
| |
|
|
|
|
| peptide tags |
minimal off-target
reactivity; operationally simple |
enzymatic conversion
efficiency is site-dependent |
deglycosylation followed
by transglutaminase treatment |
generates aglycosylated
antibody |
| transglutaminase tag |
DAR of 2 or 4 |
| sortase tag |
must be placed near C terminus |
| aldehyde tag |
conjugate stability requires
Pictet–Spengler or alternative ligation |
