An endo-1,4-beta-xylanase-encoding gene from Agaricus bisporus is regulated by compost-specific factors.
Compost is the preferred substrate for growth of the edible fungus Agaricus bisporus. Utilization of compost requires the production of enzymes involved in degradation of lignocellulolytic components. For molecular characterization of these processes we are isolating the encoding genes. By applying heterologous screening techniques, we have cloned such a gene, which is specifically induced on compost encoding an endo-1,4-beta-xylanase (xlnA) belonging to glycosyl hydrolase family 10. The gene encodes a pre-protein of 333 amino acid residues with a predicted molecular mass of 34,946 for the mature protein. The open reading frame is interrupted by ten introns of which introns 5 and 6 are separated by an exon of only two base-pairs. High expression of the xlnA gene was observed in vegetative mycelium grown on sterilized compost while xlnA messengers were not detected in fruit bodies. Addition of glucose or xylose to compost repressed xlnA expression. When glucose-grown colonies were transferred to a medium containing cellulose, xylan or xylose as sole carbon source, the organism responded by expressing xlnA at a high level for a short period. Transfer from glucose to compost yielded a much stronger and constant xlnA induction. A similar pattern of expression was found for the cel3 gene encoding a cellulase, suggesting that these genes are induced by compost-specific factors rather than by the substrates they act upon. Antiserum raised against XLNA protein, which was heterologously expressed in Escherichia coli, detected, when the fungus was grown on compost, an extracellular protein of 33 kDa with endo-xylanase activity.
De Groot
P W
PW
Section Molecular Genetics of Industrial Microorganisms, Wageningen Agricultural University, Wageningen, Dreijenlaan 2, NL-6703 HA, The Netherlands.
Basten
D E
DE
Sonnenberg
A
A
Van Griensven
L J
LJ
Visser
J
J
Schaap
P J
PJ
D000364:Agaricus / Q000201:enzymology / Q000235:genetics*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D015153:Blotting, Western; D002480:Cellulase / Q000235:genetics; D002482:Cellulose / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary; D043364:Endo-1,4-beta Xylanases; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D005947:Glucose / Q000378:metabolism; D006868:Hydrolysis; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014995:Xylosidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D018076:DNA, Complementary; D002482:Cellulose; D014995:Xylosidases; D002480:Cellulase; C061269:cel-3 protein, Fibrobacter succinogenes; D043364:Endo-1,4-beta Xylanases; D005947:Glucose