A single mutation in cyclodextrin glycosyltransferase from Paenibacillus barengoltzii changes cyclodextrin and maltooligosaccharides production

CAMINATA LANDRIEL, S; BERENGUER, J; COSTA, H; SÁNCHEZ COSTA, M; HIDALGO, A; CASTILLO, JDLM; TABOGA, O A; FERRAROTTI, S A; CAMINATA LANDRIEL, S; BERENGUER, J; COSTA, H; SÁNCHEZ COSTA, M; HIDALGO, A; CASTILLO, JDLM; TABOGA, O A; FERRAROTTI, S A

PROTEIN ENGINEERING DESIGN & SELECTION

OXFORD UNIV PRESS

Año: 2018 p. 1 - 9

1741-0126

Cyclodextrin glycosyltransferases (CGTases) are bacterial enzymes that catalyze starch conversioninto cyclodextrins, which have several biotechnological applications including solubilization ofhydrophobic compounds, masking of unpleasant odors and flavors in pharmaceutical preparations,and removal of cholesterol from food. Additionally, CGTases produce maltooligosaccharides,which are linear molecules with potential benefits for human health. Current research effortsare concentrated in the development of engineered enzymes with improved yield and/or particularproduct specificity. In this work, we analyzed the role of four residues of the CGTase fromPaenibacillus barengoltzii as determinants of product specificity. Single mutations were introducedin the CGTase-encoding gene to obtain mutants A137V, A144V, L280A and M329I and theactivity of recombinant proteins was evaluated. The residue at position 137 proved to be relevantfor CGTase activity. Molecular dynamics studies demonstrated additionally that mutation A137Vproduces a perturbation in the catalytic site of the CGTase, which correlates with a 10-fold reductionin its catalytic efficiency. Moreover, this mutant showed increased production of maltooligosaccharideswith a high degree of polymerization, mostly maltopentaose to maltoheptaose. Ourresults highlight the role of residue 137 as a determinant of product specificity in this CGTase andmay be applied to the rational design of saccharide-producing enzymes.