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Differences in the thermostabilities of barley (1→3,1→4)‐β‐glucanases are only partly determined by N ‐glycosylation
Author(s) -
Doan Danny N.P.,
Fincher Geoffrey B.
Publication year - 1992
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(92)80786-g
Subject(s) - thermostability , isozyme , glycosylation , biochemistry , chemistry , carbohydrate , enzyme , site directed mutagenesis , fructose , biology , mutant , gene
Barley (1→3,1→4)‐β‐glucan 4‐glucanohydrolase (EC 3.2.1.73) isoenzyme EII carries 4% by weight carbohydrate and is more stable at elevated temperatures than isoenzyme EI, which has no associated carbohydrate. The relationship between carbohydrate content and thermostability has been investigated by treatment of the two isoenzymes with N ‐glycopeptidase F (EC 3.5.1.52). Removal of carbohydrate from isoenzyme EII results in a decrease in the enzyme's thermostability, but treatment of isoenzyme EI with the N ‐glycopeptidase F has no effect. In addition, removal of a single N ‐glycosylation site in isoenzyme EII (Asn 190 ‐Ala‐Ser) by site‐directed mutagenesis of the corresponding cDNA led to a reduction in thermostability, while the introduction of this site into isoenzyme EI enhanced stability. We conclude that N ‐glycosylation of Asn 190 enhances the stability of isoenzyme EII at elevated temperatures, but that other factors related to their primary structures also contribute to the differences in thermostabilities of the barley (1→3,1→4)‐β‐glucanases.