aguA, the gene encoding an extracellular alpha-glucuronidase from Aspergillus tubingensis, is specifically induced on xylose and not on glucuronic acid.
An extracellular alpha-glucuronidase was purified and characterized from a commercial Aspergillus preparation and from culture filtrate of Aspergillus tubingensis. The enzyme has a molecular mass of 107 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 112 kDa as determined by mass spectrometry, has a determined pI just below 5.2, and is stable at pH 6.0 for prolonged times. The pH optimum for the enzyme is between 4.5 and 6.0, and the temperature optimum is 70 degrees C. The alpha-glucuronidase is active mainly on small substituted xylo-oligomers but is also able to release a small amount of 4-O-methylglucuronic acid from birchwood xylan. The enzyme acts synergistically with endoxylanases and beta-xylosidase in the hydrolysis of xylan. The enzyme is N glycosylated and contains 14 putative N-glycosylation sites. The gene encoding this alpha-glucuronidase (aguA) was cloned from A. tubingensis. It consists of an open reading frame of 2,523 bp and contains no introns. The gene codes for a protein of 841 amino acids, containing a eukaryotic signal sequence of 20 amino acids. The mature protein has a predicted molecular mass of 91,790 Da and a calculated pI of 5.13. Multiple copies of the gene were introduced in A. tubingensis, and expression was studied in a highly overproducing transformant. The aguA gene was expressed on xylose, xylobiose, and xylan, similarly to genes encoding endoxylanases, suggesting a coordinate regulation of expression of xylanases and alpha-glucuronidase. Glucuronic acid did not induce the expression of aguA and also did not modulate the expression on xylose. Addition of glucose prevented expression of aguA on xylan but only reduced the expression on xylose.
de Vries
R P
RP
Molecular Genetics of Industrial Microorganisms, Wageningen Agricultural University, The Netherlands.
Poulsen
C H
CH
Madrid
S
S
Visser
J
J
D000595:Amino Acid Sequence; D001230:Aspergillus / Q000201:enzymology*; D001483:Base Sequence; D003001:Cloning, Molecular; D004271:DNA, Fungal; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D005965:Glucuronates / Q000378:metabolism*; D020723:Glucuronic Acid; D006026:Glycoside Hydrolases / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D014994:Xylose / Q000378:metabolism*
D004271:DNA, Fungal; D005965:Glucuronates; D020723:Glucuronic Acid; D014994:Xylose; D006026:Glycoside Hydrolases; C096734:alpha-glucuronidase