3WWG

Crystal structure of the N-glycan-deficient variant N448A of isopullulanase complexed with isopanose

Summary for 3WWG

• Entry DOI: 10.2210/pdb3wwg/pdb
• Related: 1WMR 1X0C 2Z8G
• Related PRD ID: PRD_900001
• Descriptor: Isopullulanase, alpha-D-glucopyranose-(1-6)-beta-D-glucopyranose, alpha-D-glucopyranose-(1-4)-alpha-D-glucopyranose, ... (5 entities in total)
• Functional Keywords: beta-helix, glycoside hydrolase family 49, hydrolase
• Biological source: Aspergillus niger
• Cellular location: Secreted: O00105
• Total number of polymer chains: 4
• Total formula weight: 247746.03

Authors

Miyazaki, T.,Yashiro, H.,Nishikawa, A.,Tonozuka, T. (deposition date: 2014-06-17, release date: 2014-11-12, Last modification date: 2024-11-06)

Primary citation

Miyazaki, T.,Yashiro, H.,Nishikawa, A.,Tonozuka, T.
The side chain of a glycosylated asparagine residue is important for the stability of isopullulanase
J.Biochem., 157:225-234, 2015

PubMed Abstract: N-glycosylation has been shown to be important for the stability of some glycoproteins. Isopullulanase (IPU), a polysaccharide-hydrolyzing enzyme, is a highly N-glycosylated protein, and IPU deglycosylation results in a decrease in thermostability. To investigate the function of N-glycan in IPU, we focused on an N-glycosylated residue located in the vicinity of the active site, Asn448. The thermostabilities of three IPU variants, Y440A, N448A and S450A, were 0.5-8.4°C lower than the wild-type enzyme. The crystal structure of endoglycosidase H (Endo H)-treated N448A variant was determined. There are four IPU molecules, Mol-A, B, C and D, in the asymmetric unit. The conformation of a loop composed of amino acid residues 435-455 in Mol-C was identical to wild-type IPU, whereas the conformations of this loop in Mol-A, Mol-B and Mol-D were different from each other. These results suggest that the Asn448 side chain is primarily important for the stability of IPU. Our results indicate that mutation of only N-glycosylated Asn residue may lead to incorrect conclusion for the evaluation of the function of N-glycan. Usually, the structures of N-glycosylation sites form an extended configuration in IPU; however, the Asn448 site had an atypical structure that lacked this configuration.

PubMed: 25359784
DOI: 10.1093/jb/mvu065

Experimental method

X-RAY DIFFRACTION (2.2 Å)