 |
PDBsum entry 2vcc
|
|
|
|
References listed in PDB file
|
 |
|
Key reference
|
|
|
Title
|
|
Structural and mechanistic insight into the basis of mucopolysaccharidosis iiib.
|
|
|
Authors
|
|
E.Ficko-Blean,
K.A.Stubbs,
O.Nemirovsky,
D.J.Vocadlo,
A.B.Boraston.
|
|
|
Ref.
|
|
Proc Natl Acad Sci U S A, 2008,
105,
6560-6565.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
|
|
|
|
Abstract
|
|
|
Mucopolysaccharidosis III (MPS III) has four forms (A-D) that result from
buildup of an improperly degraded glycosaminoglycan in lysosomes. MPS IIIB is
attributable to the decreased activity of a lysosomal
alpha-N-acetylglucosaminidase (NAGLU). Here, we describe the structure,
catalytic mechanism, and inhibition of CpGH89 from Clostridium perfringens, a
close bacterial homolog of NAGLU. The structure enables the generation of a
homology model of NAGLU, an enzyme that has resisted structural studies despite
having been studied for >20 years. This model reveals which mutations giving
rise to MPS IIIB map to the active site and which map to regions distant from
the active site. The identification of potent inhibitors of CpGH89 and the
structures of these inhibitors in complex with the enzyme suggest small-molecule
candidates for use as chemical chaperones. These studies therefore illuminate
the genetic basis of MPS IIIB, provide a clear biochemical rationale for the
necessary sequential action of heparan-degrading enzymes, and open the door to
the design and optimization of chemical chaperones for treating MPS IIIB.
|
|
|
|
|
|
|
|
Figure 2.
Structural location of naturally occurring mutations in
NAGLU. (A) A cartoon representation of the homology model of
NAGLU showing its overall fold. The coloring of the domains is
as for CpGH89 in Fig. 1A. (B) A structural overlay of the active
site of CpGH89 and NAGLU. The catalytic residues are labeled.
(C) A view of the ribbon trace of the NAGLU model with the
structural location of the known mutations that lead to MPS
IIIB. Sites of mutations are shown as spheres. Blue coloring
indicates active site residues, whereas red coloring indicates
nonactive site residues.
|
|
Figure 3.
Inhibitor binding by CpGH89. (A and B) Isotherms of CpGH89
binding to PUGNAc (A) and 2AcDNJ (B) produced by ITC. (Upper)
Raw heat measurements. (Lower) Integrated heats. The solid lines
show the fit of a one site-binding model to the data. (A Inset)
Dixon plot analysis of CpGH89 inhibition by PUGNAc. The
intersection point on the graph corresponding to the K [i]
(absolute value of the X value at the intersection) is indicated
by an arrow. (C and D ) Active site representations are shown
for PUGNAc (C ) and 2AcDNJ (D). The blue mesh shows the maximum
likelihood (43)/σ[a] (49)-weighted electron density maps
contoured at 0.23 e^−/Å^3 and 0.22 e^−/Å^3 for
2AcDNJ and PUGNAc, respectively. Key active site residues,
including the putative catalytic residues Glu-483 and Glu-601,
are shown in stick representation and colored gray. Ligands are
shown in green stick representation. Putative hydrogen bonds
between the protein and ligand identified by using the criteria
of proper geometry and a distance cutoff of 3.2 Å are
shown as dotted magenta lines. (E and F ) Schematics showing the
interactions within the active site of CpGH89 with PUGNAc (E )
and 2AcDNJ (F ). A distance of 3.2 Å was used as the
cutoff for significant hydrogen bonds. Water molecules are shown
as shaded spheres. Protons on the amino acids are omitted for
clarity.
|
|
|
|
|
|
|
|
|
|
