Engineering of cyclodextrin glucanotransferases and the impact for biotechnological applications

Abstract

Cyclodextrin glucanotransferases (CGTases) are industrially important enzymes that produce cyclic α-(1,4)-linked oligosaccharides (cyclodextrins) from starch. Cyclodextrin glucanotransferases are also applied as catalysts in the synthesis of glycosylated molecules and can act as antistaling agents in the baking industry. To improve the performance of CGTases in these various applications, protein engineers are screening for CGTase variants with higher product yields, improved CD size specificity, etc. In this review, we focus on the strategies employed in obtaining CGTases with new or enhanced enzymatic capabilities by searching for new enzymes and improving existing enzymatic activities via protein engineering.

Excretory overexpression of Paenibacillus pabuli US132 cyclodextrin glucanotransferase (CGTase) in Escherichia coli: gene cloning and optimization of the culture conditions using experimental design

“…The major activities of these enzymes are intramolecular (cyclization) and intermolecular (disproportion, coupling) transglycosylation (Kuo et al 2009). They are able to convert starch into cyclic α-1,4-glucans, called cyclodextrins (CDs) (Leemhuis et al 2010). CDs can accommodate various organic and inorganic molecules within the hydrophobic central cavity leading to changes in the chemical and physical properties of the guest molecules (Li et al 2010).…”

Protein Engineering Methods and Applications

“…Screening organic solvent-tolerant bacteria or extremophiles has been preferred to isolate and improve naturally solvent-stable enzymes (Gupta & Khare, 2009;Doukyu & Ogino, 2010). Other protein engineering examples with industrially and/or pharmacologically important enzymes include studies on cholesterol oxidase (Pollegioni et al, 2009), cyclodextrin glucanotransferases (Leemhuis et al, 2010), human butyrylcholinesterase (Masson et al, 2009), microbial glucoamylases (Kumar & Satyanarayana, 2009), lipases of different origins (Akoh et al, 2004;Verma et al, 2008;Kurtovic et al, 2009), phospholipases (Song et al, 2005;De Maria et al, 2007;Simockova & Griac, 2009) and phytases (Rao et al, 2009). Studies on extremozymes, enzymes isolated from extremophilic species, revealed their different structural and functional characteristics which could be exploited for biotechnological applications and improved further by protein engineering (Bjarnason et al, 1993;Hough & Danson, 1999;Georlette et al, 2004).…”

Production of a novel Cyclodextrin glycosyltransferase from Bacillus sp. SK13.002

“…They are extracellular induced enzymes produced by microbial cells, predominantly Bacillus species. However, production by a variety of other bacterial species has also been reported (Leemhuis et al, 2010). CGTase catalyzes mainly transglycosylation reactions (cyclization, coupling and disproportionation) but can also exhibit, to a lesser extent, α-amylase-like activity, hydrolyzing starch into short linear saccharides.…”

“…There are many reports on the purification of CGTase enzymes, with most strategies used mainly involving adsorption of CGTase on starch followed by gel filtration. At present, CGTases from at least 50 host organisms have been identified and (partly) characterized, even though the genes for some have not been identified and sequenced yet (Leemhuis et al, 2010;Ramli et al, 2011). Most of the reported CGTases are monomeric, with those from Bacillus sp.…”