A type B feruloyl esterase from Aspergillus nidulans with broad pH applicability.
A hypothetical protein AN1772.2 of Aspergillus nidulans was found to have a 56% identity with a known type C ferulic acid esterase (FAE) from Talaromyces stipitatus. In addition, it contained a 13-amino acid conserved region flanking the characteristic G-X-S-X-G motif of a serine esterase, suggesting a FAE function for the protein. The putative FAE was successfully cloned from the genomic DNA and expressed in Saccharomyces cerevisiae. The recombinant protein exhibited high FAE activities. Therefore, its function as an FAE was unequivocally determined. About 86% of the enzyme activity was found in the growth medium, indicating that the native signal peptide was effective in the yeast expression system. The recombinant FAE was purified to its homogeneity, and subsequently characterized. The FAE is stable over an unusually wide range of pH (4.0-9.5), has a pH optimum of 7.0, and a temperature optimum of 45 degrees C. A substrate specificity profiling reveals that the enzyme is a type B FAE, despite its strong sequence homology with type C FAEs, raising an interesting question on the role of the conserved region in substrate specificity.
Shin
Hyun-Dong
HD
School of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, GA 30332-0100, USA.
Chen
Rachel Ruizhen
RR
D000595:Amino Acid Sequence; D001233:Aspergillus nidulans / Q000201:enzymology* / Q000235:genetics; D002265:Carboxylic Ester Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003373:Coumaric Acids / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005822:Genetic Vectors / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D013696:Temperature
D003373:Coumaric Acids; D011994:Recombinant Proteins; C004999:ferulic acid; D002265:Carboxylic Ester Hydrolases; C070196:feruloyl esterase