Purification and Characterization of Two Different -L-Rhamnosidases,RhaA and RhaB, from Aspergillus aculeatus
Two proteins exhibiting -L-rhamnosidase activity, RhaA and RhaB, were identified upon fractionation and purification of a culture filtrate from Aspergillus aculeatus grown on hesperidin. Both proteins were shown to be N glycosylated and had molecular masses of 92 and 85 kDa, of which approximately 24 and 15%, respectively, were contributed by carbohydrate. RhaA and RhaB, optimally active at pH 4.5 to 5, showed Km and Vmax values of 2.8 mM and 24 U/mg (RhaA) and 0.30 mM and 14 U/mg (RhaB) when tested for p-nitrophenyl--L-rhamnopyranoside. Both enzymes were able to hydrolyze -1,2 and -1,6 linkages to -D-glucosides. Using polyclonal antibodies, the corresponding cDNA of both -L-rhamnosidases, rhaA and rhaB, was cloned. On the basis of the amino acid sequences derived from the cDNA clones, both proteins are highly homologous (60% identity).
Main Authors: | Manzanares, Paloma, van den Broeck, Hetty C., Graaff, Leo H. de, Visser, Jaap |
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Format: | artículo biblioteca |
Language: | English |
Published: |
American Society for Microbiology
2001-05
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Subjects: | Aspergillus aculeatus, Rhamnosidases, |
Online Access: | http://hdl.handle.net/2431/73 http://hdl.handle.net/10261/3070 |
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