Biophysical Society Thematic Meeting

Biophysics in the Understanding, Diagnosis, and Treatment of Infectious Diseases Poster Abstracts

42-POS

Board 42

Enzymatic Manipulation of Antibody Fc-Mediated Effector Functions

Eric Sundberg

University of Maryland School of Medicine, Baltimore, USA.

In order to evade host immune mechanisms, many bacteria secrete a diversity of

immunomodulatory enzymes. Streptococcus pyogenes, one of the most common human

pathogens, secretes a large endoglycosidase, EndoS, which removes carbohydrates in a highly

specific manner from IgG antibodies. This renders antibodies incapable of eliciting host effector

functions through either the complement pathway or via Fc γ receptor signaling, providing the

bacteria with a survival advantage. On account of this antibody-specific modifying activity,

EndoS is currently being developed as a promising injectable therapeutic for autoimmune

diseases that rely on autoantibodies. Additionally, EndoS is a key enzyme used in the

chemoenzymatic synthesis of homogenously glycosylated antibodies with tailored Fc γ receptor-

mediated effector functions. Despite the tremendous utility of this enzyme, the molecular basis

of EndoS specificity for, and processing of, IgG antibodies has remained poorly understood. We

have recently determined the high-resolution X-ray crystal structure of EndoS, which provides

the first mechanistic insight into its unique enzymatic properties. Based on this structure, we

rationally designed chimeric endoglycosidases in which we exchanged the glycosidase domain

of EndoS with that of EndoF1 in order to create enzymes that exhibit high specificity for

antibody substrates while changing their glycan specificity to that of EndoF1 (high mannose

type) from that of EndoS (complex biantennary type). Using mass spectrometry and surface

plasmon resonance assays, we found these engineered enzymes to be highly specific and

efficient for the glycoprotein substrates for which we designed them. This novel glycoprotein

engineering strategy for constructing chimeric endoglycosidases that are able to manipulate the

glycan composition on IgG antibodies provides new opportunities to engineer antibodies with

unique glycan compositions for previously unachievable therapeutic applications.