Chemical and Structural Analysis of an Antibody Folding Intermediate Trapped during Glycan Biosynthesis

Human IgG Fc glycosylation modulates immunological effector
functions
such as antibody-dependent cellular cytotoxicity and phagocytosis.
Engineering of Fc glycans therefore enables fine-tuning of the therapeutic
properties of monoclonal antibodies. The N-linked glycans of Fc are
typically complex-type, forming a network of noncovalent interactions
along the protein surface of the Cγ2 domain. Here, we manipulate
the mammalian glycan-processing pathway to trap IgG1 Fc at sequential
stages of maturation, from oligomannose- to hybrid- to complex-type
glycans, and show that the Fc is structurally stabilized following
the transition of glycans from their hybrid- to complex-type state.
X-ray crystallographic analysis of this hybrid-type intermediate reveals
that N-linked glycans undergo conformational changes upon maturation,
including a flip within the trimannosyl core. Our crystal structure
of this intermediate reveals a molecular basis for antibody biogenesis
and provides a template for the structure-guided engineering of the
protein–glycan interface of therapeutic antibodies.