PDBsum entry 5d6t

Hydrolase

PDB id: 5d6t

Contents

Protein chain

253 a.a.

Ligands

SO4

NGA

Metals

_CL

Waters ×203

PDB id:

5d6t

Name:

Hydrolase

Title:

Crystal structure of aspergillus clavatus sph3 in complex with galnac

Structure:

Spherulin-4. Chain: a. Engineered: yes

Source:

Aspergillus clavatus (strain atcc 1007 / cbs 513.65 / dsm 816 / nctc 3887 / nrrl 1). Organism_taxid: 344612. Strain: atcc 1007 / cbs 513.65 / dsm 816 / nctc 3887 / nrrl 1. Gene: acla_035820. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

1.93Å R-factor: 0.172 R-free: 0.217

Authors:

N.C.Bamford,D.J.Little,P.L.Howell

Key ref:

N.C.Bamford et al. (2015). Sph3 Is a Glycoside Hydrolase Required for the Biosynthesis of Galactosaminogalactan in Aspergillus fumigatus. J Biol Chem, 290, 27438-27450. PubMed id: 26342082 DOI: 10.1074/jbc.M115.679050

Date:

12-Aug-15 Release date: 16-Sep-15

Protein chain

A1CJQ5  (A1CJQ5_ASPCL) -  Cell surface spherulin 4-like protein from Aspergillus clavatus (strain ATCC 1007 / CBS 513.65 / DSM 816 / NCTC 3887 / NRRL 1 / QM 1276 / 107)

Seq: 251 a.a.

Struc: 253 a.a.

DOI no: 10.1074/jbc.M115.679050

J Biol Chem 290:27438-27450 (2015)

PubMed id: 26342082

Sph3 Is a Glycoside Hydrolase Required for the Biosynthesis of Galactosaminogalactan in Aspergillus fumigatus.

N.C.Bamford, B.D.Snarr, F.N.Gravelat, D.J.Little, M.J.Lee, C.A.Zacharias, J.C.Chabot, A.M.Geller, S.D.Baptista, P.Baker, H.Robinson, P.L.Howell, D.C.Sheppard.

ABSTRACT

Aspergillus fumigatus is the most virulent species within the Aspergillus genus and causes invasive infections with high mortality rates. The exopolysaccharide galactosaminogalactan (GAG) contributes to the virulence of A. fumigatus. A co-regulated five-gene cluster has been identified and proposed to encode the proteins required for GAG biosynthesis. One of these genes, sph3, is predicted to encode a protein belonging to the spherulin 4 family, a protein family with no known function. Construction of an sph3-deficient mutant demonstrated that the gene is necessary for GAG production. To determine the role of Sph3 in GAG biosynthesis, we determined the structure of Aspergillus clavatus Sph3 to 1.25 Å. The structure revealed a (β/α)8 fold, with similarities to glycoside hydrolase families 18, 27, and 84. Recombinant Sph3 displayed hydrolytic activity against both purified and cell wall-associated GAG. Structural and sequence alignments identified three conserved acidic residues, Asp-166, Glu-167, and Glu-222, that are located within the putative active site groove. In vitro and in vivo mutagenesis analysis demonstrated that all three residues are important for activity. Variants of Asp-166 yielded the greatest decrease in activity suggesting a role in catalysis. This work shows that Sph3 is a glycoside hydrolase essential for GAG production and defines a new glycoside hydrolase family, GH135.