Purification and Characterization of an Acetyl Esterase from Aspergillus niger

James Linden, Meropi Samara, Stephen Decker, Ellen Johnson, Michele Boyer, Mlklos Pecs, William Adney, Michael Himmel

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32 Scopus Citations


Optimized acetyl esterase enzyme production conditions using Aspergillus niger ATCC 10864 in 14-L fermentation jars were determined to be 33‡C, 1.5 vvm aeration, and 300 rpm agitation without pH control. The acetyl esterase was purified by precipitation in 60-80% saturation in ammonium sulfate. The pellet was applied directly to a Pharmacia high-load Phenyl Sepharose column for hydrophobic interaction chromatography and purified to homogeneity in two steps. Stability and kinetic characteristics of the acetyl esterase were determined over a pH range of 4.0-7.5 and from 4 to 45‡C. At temperatures > 25‡C, stability was superior at pH values < 5.0. The temperature activity optimum was 35‡C, and the pH optimum was 7.0. The V max was determined to be 46,700 U/mg protein, and the K m was 0.023M p-nitrophenyl acetate at pH 6.5 in 0.2 M phosphate buffer at 35‡C. The mol wt of the enzyme was 35,000 dalton by size-exclusion chromatography and SDS gel electrophoresis. The N-terminal amino acid sequence and the glycosylation composition were also determined.

Original languageAmerican English
Pages (from-to)383-393
Number of pages11
JournalApplied Biochemistry and Biotechnology - Part A Enzyme Engineering and Biotechnology
Issue number1
StatePublished - 1994

Bibliographical note

Work performed by Department of Microbiology, Colorado State University, Fort Collins, Colorado; Institute for Agricultural Chemical Technology, Technical University of Budapest, Budapest, Hungary; and the National Renewable Energy Laboratory, Golden, Colorado

NREL Publication Number

  • ACNR/JA-15888


  • Acetyl esterase
  • acetyl xylan esterase
  • Aspergillus niger
  • hemicellulose


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