Cyclodextrin (CD) has a closed ring structure consisting of D glucose residues linked through α (1,4) linkages. The most common CDs consist of six, seven, or eight glucose residues, and are named α , β and γ CD, respectively. CDs have a hydrophilic outer surface and a relatively hydrophobic internal cavity. CDs are able to form complexes with various molecules. The formation of complexes leads to changes in the chemical and physical properties of the molecule including stability and solubility; thus there have been extensive applications of CDs in the food, cosmetic, agricultural and pharmaceutical industries. 1) Cyclodextrin glucanotransferase (CGTase; EC 2.4.1.19) forms CDs from starch and related α (1,4) linked glucose polymers through intramolecular transglycosylation. 2) This enzyme belongs to glycoside hydrolase family 13 (α amylase family), based on its amino acid sequence. 3 5) Members of this family show various reaction specificities but have the following common features: a (β α)8 barrel structure and four highly conserved regions, regions I IV, in the primary sequence. The conserved regions include invariant catalytic residues, two Asp residues and one Glu residue, and other amino acids involved in transition state stabilization. 3) A number of bacteria and archaea (e.g., Bacillus, Thermoanaerobacter and Thermococcus species) have been isolated as CGTase producers. 6) Many studies on X ray crystallographic structures of CGTases complexed with substrates, products, and inhibitors have revealed that CGTase possesses a substrate binding groove that accommodates at least 7 glucose residues at the donor subsites and 3 at the acceptor subsites (subsite −7 to +3), and contains an aromatic amino acid in a dominant position at the center of the substrate binding groove. 2,7 9) As all known CGTases produce a mixture of CDs, they are subdivided into α , β and γ CGTases based on the major CD product. The majority of CGTases reported are β CGTases while α and γ CGTases are less frequent. γ CD is the most expensive CD due to the low yield of this substance from the current CGTases. Several attempts have been made to enhance the γ CD yield, but no satisfactory results have been obtained to date. 10,11) In addition to the ability of CD synthesis (cyclization reaction), CGTase catalyzes three other reactions: coupling, disproportionation and starch hydrolysis. In the coupling reaction, CD is cleaved and transferred to an acceptor substrate. In the disproportionation reaction and starch hydrolysis reaction, a linear maltooligosaccharide is cleaved and then glycosyl residue transfers to another linear substrate and water, respectively. Thus CGTase is also utilized for the synthesis of glycosides. 12,13) Cyclodextrinase (CDase; EC 3.2.1.54) hydrolyzes CDs much faster than starch and pullulan. 14) Many CDases have been isolated from various bacteria, including Alicy- Abstract: The alkaliphilic soil bacterium Bacillus clarkii 7364 was found to produce cyclodextrin glucanotransferase (CGTase), an enzyme which conver...