Crystal structure of the conserved domain of the DC lysosomal associated membrane protein: implications for the lysosomal glycocalyx

Abstract

BackgroundThe family of lysosome-associated membrane proteins (LAMP) comprises the multifunctional, ubiquitous LAMP-1 and LAMP-2, and the cell type-specific proteins DC-LAMP (LAMP-3), BAD-LAMP (UNC-46, C20orf103) and macrosialin (CD68). LAMPs have been implicated in a multitude of cellular processes, including phagocytosis, autophagy, lipid transport and aging. LAMP-2 isoform A acts as a receptor in chaperone-mediated autophagy. LAMP-2 deficiency causes the fatal Danon disease. The abundant proteins LAMP-1 and LAMP-2 are major constituents of the glycoconjugate coat present on the inside of the lysosomal membrane, the 'lysosomal glycocalyx'. The LAMP family is characterized by a conserved domain of 150 to 200 amino acids with two disulfide bonds.ResultsThe crystal structure of the conserved domain of human DC-LAMP was solved. It is the first high-resolution structure of a heavily glycosylated lysosomal membrane protein. The structure represents a novel β-prism fold formed by two β-sheets bent by β-bulges and connected by a disulfide bond. Flexible loops and a hydrophobic pocket represent possible sites of molecular interaction. Computational models of the glycosylated luminal regions of LAMP-1 and LAMP-2 indicate that the proteins adopt a compact conformation in close proximity to the lysosomal membrane. The models correspond to the thickness of the lysosomal glycoprotein coat of only 5 to 12 nm, according to electron microscopy.ConclusionThe conserved luminal domain of lysosome-associated membrane proteins forms a previously unknown β-prism fold. Insights into the structure of the lysosomal glycoprotein coat were obtained by computational models of the LAMP-1 and LAMP-2 luminal regions.

CD68/macrosialin: not just a histochemical marker

“…It is predicted that all LAMP family proteins contain LAMP domains of similar structure (Figure 2). 31,33 The DC-LAMP domain consists of two β-sheets. 33 The domain's N-and C-termini are located on the 'front' β-sheet, which consists of six β-strands.…”

“…31,33 The DC-LAMP domain consists of two β-sheets. 33 The domain's N-and C-termini are located on the 'front' β-sheet, which consists of six β-strands. The β-strands S1 and S3 are short and arranged in tandem, antiparallel to strand S2.…”

“…Two more antiparallel β-strands, S10 and S9, are followed by the short C-terminal β-strand S11, which is arranged in parallel to S9. 33 The C-terminal 10-residue tail (RRRFSAYQAL in human CD68 and RRRQSTYQPL in murine macrosialin), is highly conserved in LAMPs indicating lysosomal-specific targeting. While some lysosomal membrane proteins are delivered to lysosomes indirectly (through endocytosis), others such as CD68 reach endosomes/lysosomes primarily from the transGolgi network in clathrin-coated vesicles.…”

“…The topology of the β-sheets is drawn schematically in the lower part with sheets opened out. (This figure is reproduced from Wilke et al, 33 with permission from BioMed Central). for CD68 in inflammatory responses of innate and humoral immunity.…”

Glycosylation inhibition reduces cholesterol accumulation in NPC1 protein-deficient cells

“…The limiting membrane of lysosomes is lined predominantly by densely packed, highly glycosylated, lysosomal membrane proteins; their oligosaccharides are thought to protect the phospholipid bilayer from hydrolytic destruction (11,12). This glycoprotein coat can be visualized by electron microscopy and has an average thickness of about 8 nm (13).…”