Protein Engineering, Vol. 13, No. 7, 509-513,
July 2000
© 2000 Oxford University Press
Conversion of the maltogenic 
-amylase Novamyl into a CGTase
Novo Nordisk A/S, Novo Allé, DK-2800 Bagsvaerd, Denmark
Novamyl is a thermostable five-domain maltogenic 
-amylase that shows sequence and structural homology with the cyclodextrin glycosyltransferases (CGTases). Comparing X-ray crystal structures of Novamyl and CGTases, two major differences in the active site cleft were observed: Novamyl contains a loop insertion consisting of five residues (residues 191–195) and the location of an aromatic residue known to be essential to obtain an efficient cyclization reaction. To convert Novamyl into a cyclodextrin (CD)-producing enzyme, the loop was deleted and two substitutions, F188L and T189Y, were introduced. Unlike the parent Novamyl, the obtained variant is able to produce ß-CD and showed an overall conversion of starch to CD of 9%, compared with CGTases which are able to convert up to 40%. The lower conversion compared with the CGTase is probably due to additional differences in the active site cleft and in the starch-binding E domain. A variant with only the five-residue loop deleted was not able to form ß-CD.
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