title: Cyclodextrin glycosyltransferase: a key enzyme in the assimilation of starch by the halophilic archaeon Haloferax mediterranei.
abstract: A cyclodextrin glycosyltransferase (CGTase, EC 2.4.1.19) was successfully isolated and characterized from the halophilic archaeon Haloferax mediterranei. The enzyme is a monomer with a molecular mass of 77 kDa and optimum activity at 55°C, pH 7.5 and 1.5 M NaCl. The enzyme displayed many activities related to the degradation and transformation of starch. Cyclization was found to be the predominant activity, yielding a mixture of cyclodextrins, mainly α-CD, followed by hydrolysis and to a lesser extent coupling and disproportionation activities. Gene encoding H. mediterranei CGTase was cloned and heterologously overexpressed. Sequence analysis revealed an open reading frame of 2142 bp that encodes a protein of 713 amino acids. The amino acid sequence displayed high homology with those belonging to the α-amylase family. The CGTase is secreted to the extracellular medium by the Tat pathway. Upstream of the CGTase gene, four maltose ABC transporter genes have been sequenced (malE, malF, malG, malK). The expression of the CGTase gene yielded a fully active CGTase with similar kinetic behavior to the wild-type enzyme. The H. mediterranei CGTase is the first halophilic archaeal CGTase characterized, sequenced and expressed.
authors: División de Bioquímica y Biología Molecular, Departamento de Agroquímica y Bioquímica, Facultad de Ciencias, Universidad de Alicante, Alicante, Spain. vanesa.bautista@ua.es
authors: Vanesa
authors: V
authors: Bautista
authors: Julia
authors: J
authors: Esclapez
authors: Francisco
authors: F
authors: Pérez-Pomares
authors: Rosa María
authors: RM
authors: Martínez-Espinosa
authors: Mónica
authors: M
authors: Camacho
authors: María José
authors: MJ
authors: Bonete
mesh_terms: D002853:Chromatography, Liquid; D003500:Cyclization; D004591:Electrophoresis, Polyacrylamide Gel; D005964:Glucosyltransferases / Q000737:chemistry / Q000378:metabolism*; D019623:Haloferax mediterranei / Q000201:enzymology* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008970:Molecular Weight; D021241:Spectrometry, Mass, Electrospray Ionization; D013213:Starch / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D013213:Starch; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase