X-ray crystallographic native sulfur SAD structure determination of laminarinase Lam16A from Phanerochaete chrysosporium.
Laminarinase Lam16A from Phanerochaete chrysosporium was recombinantly expressed in Pichia pastoris, crystallized and the structure was solved at 1.34 A resolution using native sulfur SAD X-ray crystallography. It is the first structure of a non-specific 1,3(4)-beta-D-glucanase from glycoside hydrolase family 16 (GH16). P. chrysosporium is a wood-degrading basidiomycete fungus and Lam16A is the predominant extracellular protein expressed when laminarin is used as the sole carbon source. The protein folds into a curved beta-sandwich homologous to those of other known GH16 enzyme structures (especially kappa-carrageenase from Pseudoalteromonas carrageenovora and beta-agarase from Zobelia galactanivorans). A notable likeness is also evident with the related glycoside hydrolase family 7 (GH7) enzymes. A mammalian lectin, p58/ERGIC, as well as polysaccharide lyase (PL7) enzymes also showed significant similarity to Lam16A. The enzyme has two potential N-glycosylation sites. One such site, at Asn43, displayed a branched heptasaccharide sufficiently stabilized to be interpreted from the X-ray diffraction data. The other N-glycosylation motif was found close to the catalytic centre and is evidently not glycosylated.
Vasur
Jonas
J
Department of Molecular Biology, Swedish University of Agricultural Sciences, PO Box 590, SE-75124 Uppsala, Sweden.
Kawai
Rie
R
Larsson
Anna M
AM
Igarashi
Kiyohiko
K
Sandgren
Mats
M
Samejima
Masahiro
M
Ståhlberg
Jerry
J
D020134:Catalytic Domain; D018360:Crystallography, X-Ray / Q000379:methods; D044644:Endo-1,3(4)-beta-Glucanase / Q000737:chemistry* / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics; D015870:Gene Expression; D005936:Glucans; D006031:Glycosylation; D006801:Humans; D037241:Mannose-Binding Lectins / Q000737:chemistry / Q000235:genetics; D020075:Phanerochaete / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D010843:Pichia / Q000235:genetics / Q000254:growth & development; D011134:Polysaccharides / Q000494:pharmacology; D046188:Protein Modification, Translational; D017434:Protein Structure, Tertiary
D005656:Fungal Proteins; D005936:Glucans; D037241:Mannose-Binding Lectins; D011134:Polysaccharides; C008247:laminaran; D044644:Endo-1,3(4)-beta-Glucanase