Isolation and characterization of a 1,4-β-endoxylanase gene of A. awamori

Abstract

An enzyme with a particular 1,4-beta-xylanase activity was identified and purified from wheat-bran culture medium of an Aspergillus awamori strain. With oligonucleotides based on the N-terminal amino-acid sequence of the enzyme, the exlA gene of A. awamori, encoding 1,4-beta-xylanase A, has been cloned. Based on the deduced amino-acid sequence, 1,4-beta-xylanase A is produced as a 211 amino-acid-residue-long precursor, which is converted post-translationally into a 184-aa residue-long mature protein. Transformation of the original A. awamori strain with multiple copies of the exlA gene resulted in a 40-fold overproduction of 1,4-beta-xylanase A. The overproduced enzyme has the same biochemical and enzymological properties as the wild-type enzyme.

Regulation of the amylolytic and (hemi-)cellulolytic genes in aspergilli

“…In the past few years, significant progress has been made in cloning several xylanolytic genes from filamentous fungi such as Aspergillus awamori (Hessing et al, 1994), Aspergillus kawachii (Ito et al, 1992a, b), A. nidulans (Jose et al, 1996;MacCabe et al, 1996;Perez-Gonzalez et al, 1996, 1998, A. oryzae Kitamoto et al, 1998), A. niger (Kinoshita et al, 1995), A. tubingensis (de Graaff et al, 1994), Chaetomium gracile (Yoshino et al, 1995), Cochliobolus carbonum (Apel et al, 1993), Humicola isolens (Dalbøge and Heldt-Hansen, 1994), Penicillium chrysogenum (Haas et al, 1993), T. reesei (Torronen et al, 1992), and Trichoderma viride (Ujiie et al, 1991). Besides these xylanolytic genes, many genes encoding accessory enzymes such as acetylxylan esterase (A. tubingensis axeA-de Graaff et al, 1992;T.…”

A Transcriptional Activator, AoXlnR, Controls the Expression of Genes Encoding Xylanolytic Enzymes in Aspergillus oryzae

“…A number of xylanase-encoding genes have been cloned from filamentous fungi such as Aspergillus awamori (Hessing et al, 1994), Aspergillus kawachii (Ito et al, 1992a,b), Aspergillus nidulans (MacCabe et al, 1996;Perez-Gonzalez et al, 1996, Aspergillus oryzae Kimura et al, 1998), Aspergillus niger (Kinoshita et al, 1995), Aspergillus tubigensis (de Graaff et al, 1994), Chaetomium gracile (Yoshino et al, 1995), Cochliobolus carbonum (Apel et al, 1993), Humicola isolens (Dalboge and Heldt-Hansen, 1994), Penicillium chrysogenum (Haas et al, 1993), Trichoderma reesei (Torronen et al, 1992), and Trichoderma viride (Ujiie et al, 1991). Recently, a gene encoding a transcriptional activator, XlnR, involved in expression of the xylanolytic genes has been cloned from A. niger, and the XlnR binding sequence, 5Ј-GGCTAAA-3Ј, has also been identified by gel mobility shift assays, DNase I footprinting, and in vivo expression studies (van Peij et al, 1998a).…”

Specific Characterization of Substrate and Inhibitor Binding Sites of a Glycosyl Hydrolase Family 11 Xylanase fromAspergillus niger

“…Subsites Ϫ3, Ϫ2, and Ϫ1 are well characterized by the inspection of the complex structures (13,16,17), but the characterization of the aglycone subsites is based only on modeling (22,26). Aspergillus niger xylanase is a 20-kDa family 11 xylanase with a pI and a pH optimum of 3.5 (32,33) for which an x-ray crystal structure is available (24). The active site is located within a deep and long cleft, which is lined with many aromatic amino acid residues and is large enough to accommodate at least four xylose residues (24).…”