Cation-independent mannose 6-phosphate receptor: a composite of distinct phosphomannosyl binding sites

J Biol Chem. 2009 Dec 11;284(50):35215-26. doi: 10.1074/jbc.M109.056184. Epub 2009 Oct 19.

Abstract

The 300-kDa cation-independent mannose 6-phosphate receptor (CI-MPR), which contains multiple mannose 6-phosphate (Man-6-P) binding sites that map to domains 3, 5, and 9 within its 15-domain extracytoplasmic region, functions as an efficient carrier of Man-6-P-containing lysosomal enzymes. To determine the types of phosphorylated N-glycans recognized by each of the three carbohydrate binding sites of the CI-MPR, a phosphorylated glycan microarray was probed with truncated forms of the CI-MPR. Surface plasmon resonance analyses using lysosomal enzymes with defined N-glycans were performed to evaluate whether multiple domains are needed to form a stable, high affinity carbohydrate binding pocket. Like domain 3, adjacent domains increase the affinity of domain 5 for phosphomannosyl residues, with domain 5 exhibiting approximately 60-fold higher affinity for lysosomal enzymes containing the phosphodiester Man-P-GlcNAc when in the context of a construct encoding domains 5-9. In contrast, domain 9 does not require additional domains for high affinity binding. The three sites differ in their glycan specificity, with only domain 5 being capable of recognizing Man-P-GlcNAc. In addition, domain 9, unlike domains 1-3, interacts with Man(8)GlcNAc(2) and Man(9)GlcNAc(2) oligosaccharides containing a single phosphomonoester. Together, these data indicate that the assembly of three unique carbohydrate binding sites allows the CI-MPR to interact with the structurally diverse phosphorylated N-glycans it encounters on newly synthesized lysosomal enzymes.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Binding Sites
  • Carbohydrate Conformation
  • Carbohydrate Sequence
  • Glucuronidase / genetics
  • Glucuronidase / metabolism
  • Humans
  • Mannosephosphates / chemistry
  • Mannosephosphates / metabolism*
  • Microarray Analysis
  • Molecular Sequence Data
  • Phosphorylation
  • Polysaccharides / analysis
  • Receptor, IGF Type 2 / chemistry*
  • Receptor, IGF Type 2 / genetics
  • Receptor, IGF Type 2 / metabolism*

Substances

  • Mannosephosphates
  • Polysaccharides
  • Receptor, IGF Type 2
  • mannose-6-phosphate
  • Glucuronidase