Structure-based mutagenesis reveals the albumin-binding site of the neonatal Fc receptor

Abstract

Albumin is the most abundant protein in blood where it has a pivotal role as a transporter of fatty acids and drugs. Like IgG, albumin has long serum half-life, protected from degradation by pH-dependent recycling mediated by interaction with the neonatal Fc receptor, FcRn. Although the FcRn interaction with IgG is well characterized at the atomic level, its interaction with albumin is not. Here we present structure-based modelling of the FcRn–albumin complex, supported by binding analysis of site-specific mutants, providing mechanistic evidence for the presence of pH-sensitive ionic networks at the interaction interface. These networks involve conserved histidines in both FcRn and albumin domain III. Histidines also contribute to intramolecular interactions that stabilize the otherwise flexible loops at both the interacting surfaces. Molecular details of the FcRn–albumin complex may guide the development of novel albumin variants with altered serum half-life as carriers of drugs.

Structural Insights into Neonatal Fc Receptor-based Recycling Mechanisms

“…Indeed, it was reported that DIII alone exhibits significant binding to human FcRn, whereas DI and DII individually or together show no measurable interaction (6). Andersen et al (13) and Schmidt et al (14) also found that DIII plays a crucial role in HSA pH-dependent binding to FcRn, although it was also suggested that DI and DII could also bring a moderate contribution (13). Our findings agree well with these data and reaffirm the crucial role played by HSA DIII, along with a significant contribution of DI.…”

“…Such a modified humanized IgG exhibited an ϳ2-4-and 4-fold increase in its serum half-life in human and cynomolgus monkeys, respectively (11,12). Other studies with human serum albumin (HSA) used modeling and crystallographic approaches coupled with mutagenesis to identify several important HSA residues, and pinpointed the crucial role played by its domain III (13,14).…”

Interaction with Both Domain I and III of Albumin Is Required for Optimal pH-dependent Binding to the Neonatal Fc Receptor (FcRn)

“…To gain insight into how FcRn binds albumin pH dependently, we and others have previously reported on mapping of the core FcRn-albumin interaction interface using site-directed mutagenesis (14,16,18). These studies revealed that the C-terminal DIII contains the principal binding site for FcRn.…”

Hepatic FcRn regulates albumin homeostasis and susceptibility to liver injury

“…These sites were centered on several previously identified albumin contact residues in FcRn, Phe157 (F157), His161 (H161), and His166 (H166) (Fig. 4 G and H), and provide evidence for the bona-fide nature of this FcRn-albuminbinding mimic (12,43). Furthermore, comparison of the FcRn: (SYN1753) 2 complex with previously published FcRn-albumin crystallographic studies (37) showed that the pair of SYN1753 peptides bind hFcRn at the same binding site as domain I of albumin (Fig.…”

“…Although albumin and IgG display pHdependent binding to FcRn, the interaction surface, affinity, and mode of interaction differ between the two proteins (11)(12)(13). Still,…”