Twists and Turns of the Cation-dependent Mannose 6-Phosphate Receptor

Abstract

Mannose 6-phosphate receptors (MPRs) participate in the biogenesis of lysosomes in higher eukaryotes by transporting soluble acid hydrolases from the transGolgi network to late endosomal compartments. The receptors release their ligands into the acidic environment of the late endosome and then return to the transGolgi network to repeat the process. However, the mechanism that facilitates ligand binding and dissociation upon changes in pH is not known. We report the crystal structure of the extracytoplasmic domain of the homodimeric cation-dependent MPR in a ligand-free form at pH 6.5. A comparison of the ligand-bound and ligand-free structures reveals a significant change in quaternary structure as well as a reorganization of the binding pocket, with the most prominent change being the relocation of a loop (residues Glu 134 -Cys 141 ). The movements involved in the bound-to-free transition of the cation-dependent MPR are reminiscent of those of the oxy-to-deoxy hemoglobin transition. These results allow us to propose a mechanism by which the receptor regulates its ligand binding upon changes in pH; the pK a of Glu 133 appears to be responsible for ligand release in the acidic environment of the late endosomal compartment, and the pK a values of the sugar phosphate and His 105 are accountable for its inability to bind ligand at the cell surface where the pH is about 7.4. P-type lectins function as an essential part of the degradative pathway of higher eukaryotic organisms. A well characterized function of this receptor family is its ability to target newly synthesized acid hydrolases to the lysosome. During their transport through the Golgi complex, acid hydrolases acquire a mannose 6-phosphate (Man-6-P) 1 recognition marker on their N-linked oligosaccharides that serves as a high affinity ligand for the receptors. The resulting receptor-acid hydrolase complex is transported to late endosomal compartments. Upon encountering the lower pH of this prelysosomal compartment, the complex dissociates. The hydrolytic enzymes are then packaged into lysosomes, and the uncomplexed receptors move back to the Golgi where the process is repeated numerous times for a single receptor (1-3).The 46-kDa cation-dependent mannose 6-phosphate receptor (CD-MPR) and the 300-kDa insulin-like growth factor II/cation-independent mannose 6-phosphate receptor (IGFII/CI-MPR) are the two known members of the P-type lectin family, both of which are type I membrane glycoproteins. Carbohydrate-binding proteins have historically been divided into two subgroups. The group I carbohydrate-binding proteins, which include periplasmic transport proteins and some enzymes, typically envelop their ligands in deep pockets. In contrast, group II carbohydrate-binding proteins, which include lectins, typically bind their ligands in shallow clefts on the protein surface (4). Our recent crystallographic studies of the CD-MPR in complex with ligands containing phosphomannosyl residues reveal that this P-type lectin is unusual with respect to the architecture of i...

Structural Insights into the Mechanism of pH-dependent Ligand Binding and Release by the Cation-dependent Mannose 6-Phosphate Receptor

“…The quaternary structural change resulting from this scissoring and twisting motion preserves the exact molecular two-fold symmetry of the dimeric molecule. In addition, a change in the distance between the individual ligand binding sites is also observed; the C␣ atoms of His-105 located at the tip of loop C are located ϳ34 Å apart in the open conformation and ϳ26 Å apart in the closed conformation (13).…”

“…State-We have previously determined the structure of the sCD-MPR at pH 6.5 in the absence of ligand (13). To determine the effect of pH on the conformation of the unbound form of the receptor, we have now determined the structure of the receptor at pH 4.8 in the absence of ligand.…”

“…We previously reported the crystallization of the extracytoplasmic region (residues 1-154) of the CD-MPR in complex with either Man-6-P (11) or the oligosaccharide pentamannosyl phosphate (P-Man(␣1,3)Man(␣1,3)Man(␣1,3)Man(␣1,2)Man) (12). We had also obtained the structure of the CD-MPR in a ligand-free form at pH 6.5, which showed a surprising finding not observed in other lectins; a significant change in the quaternary structure as well as a relocation of a loop in the binding pocket was observed when compared with the structure of the CD-MPR in a ligand-bound state at pH 6.5 (13). However, it was unclear whether the CD-MPR in a ligand-unbound state could adopt other conformations that are dependent upon pH.…”

N-Glycosylation Is Crucial for Folding, Trafficking, and Stability of Human Tripeptidyl-peptidase I

“…Both MPRs, the 46-kDa cation-dependent MPR and the 300-kDa insulin-like growth factor II/cation-independent MPR (IG-FII/CI-MPR), participate in sorting and lysosomal targeting of acid hydrolases in trans-Golgi compartment, but only IGFII/ CI-MPR participates in endocytosis of lysosomal enzymes (68,69). Although the affinity of IGFII/CI-MPR to acid hydrolases is generally higher than that of cation-dependent MPR, they both participate in lysosomal targeting of largely overlapping complement of lysosomal enzymes, and neither receptor can fully compensate for the absence of the other (70).…”

Identification of a Low Affinity Mannose 6-Phosphate-binding Site in Domain 5 of the Cation-independent Mannose 6-Phosphate Receptor

“…Our crystal structures of the MPRs (13,(27)(28)(29)44) have provided insight into the mechanism by which these receptors recognize Man-6-P with high affinity. The core structure (Fig.…”