Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2012 Jul 19:10:62.
doi: 10.1186/1741-7007-10-62.

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

Sonja Wilke  1 , Joern Krausze, Konrad Büssow
Affiliations

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

Sonja Wilke et al. BMC Biol. .

Abstract

Background: The 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.

Results: The 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.

Conclusion: The 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.

PubMed Disclaimer

Figures

Figure 1
Figure 1
The LAMP protein family. (A) Domain architectures of the five human LAMP family proteins (UniProt P11279, P13473, Q9UQV4, Q9UJQ1, P34810). O- and N-linked glycosylation sites are indicated in blue and red, respectively. SP, signal peptide; TM, transmembrane helix. (B, C) Sequence alignments of the seven human LAMP domains and adjacent transmembrane domains (B) and of the 'hinge' region (linker) connecting the membrane-distal ('dist') and membrane-proximal ('prox') domains of LAMP-1 and LAMP-2 (C). Glycosylated residues are displayed with yellow background. Residues involved in β-bulges are indicated by arcs in magenta. Hydrophobic residues are colored red and hydrophilic ones are colored blue (in B). S-S: disulfide bond.
Figure 2
Figure 2
The structure of the DC-LAMP domain. (A) Different views of the domain's shape. The two β-sheets are drawn in red and blue. (B) Stereo views of the domain's β-prism shape. Schematic prism shapes are drawn on the right for orientation. The N-acetyl-glucosamine residue is depicted as a stick drawing in cyan.
Figure 3
Figure 3
The β-strand arrangement of the DC-LAMP domain. The 'front' and 'back' β-sheets are drawn in red and blue, respectively. The cysteines that form the first disulfide bond (S-S) are labelled. β-strands and loops are identified as S1 to S11 and L1 to L3, respectively. The topology of the β-sheets is drawn schematically in the lower part with sheets opened out.
Figure 4
Figure 4
Stereo views of the β-bulges. The β-bulges that bend the front and back β-sheets are shown. Main chain atoms of parts of β-strands are shown as stick drawings. Bulged residues are numbered and their conformations are given in parentheses (α, α-helix; β, β-strand; I', type I' β-turn; Lα, left-handed α-helix). Hydrogen bond pairs that flank β-bulges are shown in red and the corresponding residues are labelled. Other inter-main chain hydrogen bonds are drawn in grey.
Figure 5
Figure 5
The hydrophobic pocket and the variable loop conformation. (A) The hydrophobic pocket in the DC-LAMP domain was filled with water molecules, which are displayed as a red surface, using HOLLOW [70]. (B) Structural alignment of DC-LAMP chain A (grey) and B (blue). Hydrogen bonds connect loop L1 to an adjacent hairpin in chain A, but not in chain B. (C) A VH immunoglobulin domain is shown for comparison (PDB 1IGT). Complementarity determining regions (CDR) are drawn as red loops. The two β-sheets of the domain, displayed in green and blue, are connected by a disulfide bond (S-S).
Figure 6
Figure 6
Model of lysosomal membrane proteins and the glycoprotein coat. Structural models of glycosylated DC-LAMP and LAMP-1 were drawn to scale. The models are based on the DC-LAMP crystal structure and a hypothetical in silico model of LAMP-1. The thickness of the glycoprotein coat was reported to range from 5 to 12 nm with an average of 8 nm. The membrane-distal, N-terminal domain of LAMP-1 may adopt other positions. Dotted lines indicate putative binding sites consisting of the flexible loop L1 and an adjacent β-hairpin (Figure 5B). A box corresponding to 120 nm2 surface area was drawn around a top view of LAMP-1. Polypeptides are depicted in green and glycans in yellow.
Figure 7
Figure 7
Molecular modeling. (A) The structure of the lowest energy obtained by molecular dynamics simulation is shown as a stick model and a van der Waals surface with pink carbon atoms. Glycans are shown with yellow carbon atoms. Prolines 203 and 206 have α-helical conformations. (B) Radius of gyration values during the simulation and r.m.s. deviations of the backbone atoms from the structure of lowest energy (marked in red). The peptide's lowest-energy structure had a radius of gyration of 14.6 Å, compared to 17.9 Å for the initial structure.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Braulke T, Bonifacino JS. Sorting of lysosomal proteins. Biochim Biophys Acta. 2009;1793:605–614. doi: 10.1016/j.bbamcr.2008.10.016. - DOI - PubMed
    1. Huotari J, Helenius A. Endosome maturation. Embo J. 2011;30:3481–3500. doi: 10.1038/emboj.2011.286. - DOI - PMC - PubMed
    1. Pillay CS, Elliott E, Dennison C. Endolysosomal proteolysis and its regulation. Biochem J. 2002;363:417–429. doi: 10.1042/0264-6021:3630417. - DOI - PMC - PubMed
    1. Luzio JP, Pryor PR, Bright NA. Lysosomes: fusion and function. Nat Rev Mol Cell Biol. 2007;8:622–632. doi: 10.1038/nrm2217. - DOI - PubMed
    1. Bright NA, Gratian MJ, Luzio JP. Endocytic delivery to lysosomes mediated by concurrent fusion and kissing events in living cells. Curr Biol. 2005;15:360–365. doi: 10.1016/j.cub.2005.01.049. - DOI - PubMed

Publication types

MeSH terms

Substances

Associated data