title: Lectin-glycoenzyme column chromatography monitored by enzyme flow microcalorimetry.
abstract: A method based on the flow microcalorimetric determination of catalytic activity of immobilized enzyme in a so-called enzyme thermistor was used to monitor the process of lectin affinity chromatography of invertase on Concanavalin A-bead cellulose. The strong biospecific interaction between Concanavalin A and invertase was employed to determine the bound enzyme and this principle was used for the investigation of an alternative direct method for monitoring the lectin affinity chromatography of glycoenzymes. The results obtained by flow microcalorimetry showed that the catalytic activity of invertase immobilized on Concanavalin A-bead cellulose can be compared directly with the thermometric value delta Tmax. The validity of the method was also confirmed by the enzyme thermistor post-column method, which is based on the determination of the product from the immobilized invertase enzymatic reaction. The adsorption and desorption in the chromatography column were examined by flow microcalorimetry in small samples withdrawn from the column. Attention has been given to the operating parameters and the storage stability of the affinity sorbent. The binding ability of the affinity matrix decreased with the number of consecutive chromatographic runs, although its storage stability was satisfactory.
authors.affiliation: Institute of Chemistry, Slovak Academy of Sciences, Slovak Republic.
authors.forename: D
authors.initials: D
authors.lastname: Mislovicová
authors.forename: M
authors.initials: M
authors.lastname: Chudinová
authors.forename: A
authors.initials: A
authors.lastname: Vikartovská
authors.forename: P
authors.initials: P
authors.lastname: Gemeiner
mesh_terms: D002138:Calibration; D002151:Calorimetry; D002846:Chromatography, Affinity; D003208:Concanavalin A; D004800:Enzymes, Immobilized; D006023:Glycoproteins / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D037102:Lectins / Q000302:isolation & purification*
chemical_list: D004800:Enzymes, Immobilized; D006023:Glycoproteins; D037102:Lectins; D003208:Concanavalin A