Purification and characterization of a chlorogenic acid hydrolase from Aspergillus niger catalysing the hydrolysis of chlorogenic acid

• Published in: Journal of biotechnology 2005-01, Vol.115 (1), p.47-56
• Main Authors: Asther, Michèle, Estrada Alvarado, Maria Isabel, Haon, Mireille, Navarro, David, Asther, Marcel, Lesage-Meessen, Laurence, Record, Eric
• Format: Article
• Language: English
• Subjects: Aspergillus niger; Aspergillus niger - classification; Aspergillus niger - enzymology; Biological and medical sciences; Biotechnology; Caffeic acid; Caffeic Acids - chemical synthesis; Carboxylic Ester Hydrolases - chemistry; Carboxylic Ester Hydrolases - isolation & purification; Catalysis; Chemical and Process Engineering; Chlorogenic acid; Chlorogenic Acid - chemistry; Chlorogenic acid hydrolase; Cinnamoyl ester hydrolase; Engineering Sciences; Enzyme Activation; Enzyme Stability; Food engineering; Fundamental and applied biological sciences. Psychology; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Life Sciences; Molecular Weight; Species Specificity; Substrate Specificity; Temperature
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/j.jbiotec.2004.07.009

Among 15 Aspergillus strains, Aspergillus niger BRFM 131 was selected for its high chlorogenic acid hydrolase activity. The enzyme was purified and characterized with respect to its physico-chemical and kinetic properties. Four chromatographic steps were necessary to purify the protein to homogeneity with a recovery of 2%. K m of the chlorogenic acid hydrolase was estimated to be 10 μM against chlorogenic acid as substrate. Under native conditions, the protein presented a molecular mass of 170 kDa, and SDS-PAGE analysis suggested the presence of two identical 80 kDa subunits. Isoelectric point was 6.0; pH optimum for activity was determined to be 6.0 and temperature optima to be 55 °C. The N-terminal sequence did not present any homology with other cinnamoyl ester hydrolases previously described suggesting the purification of a new protein. The chlorogenic acid hydrolase was used successfully for the production of caffeic acid, which possesses strong antioxidant properties, from natural substrates specially rich in chlorogenic acid like apple marc and coffee pulp.