Structural Characterization of Carbohydrate Binding by LMAN1 Protein Provides New Insight into the Endoplasmic Reticulum Export of Factors V (FV) and VIII (FVIII)*

LMAN1 (ERGIC-53) is a key mammalian cargo receptor responsible for the export of a subset of glycoproteins from the endoplasmic reticulum. Together with its soluble coreceptor MCFD2, LMAN1 transports coagulation factors V (FV) and VIII (FVIII). Mutations in LMAN1 or MCFD2 cause the genetic bleeding disorder combined deficiency of FV and FVIII (F5F8D). The LMAN1 carbohydrate recognition domain (CRD) binds to both glycoprotein cargo and MCFD2 in a Ca²⁺-dependent manner. To understand the biochemical basis and regulation of LMAN1 binding to glycoprotein cargo, we solved crystal structures of the LMAN1-CRD bound to Man-α-1,2-Man, the terminal carbohydrate moiety of high mannose glycans. Our structural data, combined with mutagenesis and in vitro binding assays, define the central mannose-binding site on LMAN1 and pinpoint histidine 178 and glycines 251/252 as critical residues for FV/FVIII binding. We also show that mannobiose binding is relatively independent of pH in the range relevant for endoplasmic reticulum-to-Golgi traffic, but is sensitive to lowered Ca²⁺ concentrations. The distinct LMAN1/MCFD2 interaction is maintained at these lowered Ca²⁺ concentrations. Our results suggest that compartmental changes in Ca²⁺ concentration regulate glycoprotein cargo binding and release from the LMAN1·MCFD2 complex in the early secretory pathway.

Background: LMAN1 is an important mammalian cargo receptor for endoplasmic reticulum (ER)-to-Golgi trafficking.

Results: Crystal structures pinpoint critical residues on LMAN1 for mannose binding, which is sensitive to Ca²⁺ concentration.

Conclusion: Changes in Ca²⁺ concentration can cause ligand release without disrupting the LMAN1·MCFD2 receptor complex.

Significance: The results shed new light on how LMAN1 binds and releases its cargo in the ER-to-Golgi transport pathway.