Purification and characterization of a new xylanase (xylanase C) produced by Streptomyces lividans 66 |
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Authors: | Dieter Kluepfel Nicole Daigneault Rolf Morosoli François Shareck |
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Institution: | (1) Centre de Recherche en Microbiologie Appliquée, Institut Armand-Frappier, Université du Québec, 531 boulevard des Prairies, H7N 4Z3 Ville de Laval, Québec, Canada |
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Abstract: | Summary A third extracellular xylanase produced by Streptomyces lividans 66 was isolated from a clone obtained by shotgun cloning through functional complementation of a xylanase- and cellulase-negative mutant using the multicopy vector pIJ702. This enzyme, designated xylanase C, has a relative molecular mass of 22000 and acts on xylan similarly to xylanase B as an endo-type xylanase producing short-chain oligoxylosides. Its specific activity determined at 1100 IU·mg–1 of protein corresponds on a molecular basis to that of xylanase B and is about three times that of xylanase A. The enzyme shows optimal activity at pH 6.0 and 57°C, values that correspond closely to those observed previously for xylanase A and B. Xylanase C appears not to be glycosylated and has a pI > 10.25. Its K
m and V
max on birchwood xylan are 4.1 mg·ml–1 and 3.0 mol·min–1·mg–1 of enzyme respectively. Whereas specific antibodies raised against xylanase A show no cross-reaction with either xylanase B or with xylanase C, the anti-(xylanase C) antibodies react slightly with xylanase B but not with xylanase A. A comparison of hydrolysis products obtained by reacting individually the three enzymes with birchwood xylan showed characteristic endo-activity patterns for xylanases B and C, whereas xylanase A hydrolysed the substrate preferentially into xylobiose and xylotriose. Sequential xylanase action on the same substrates showed synergistic hydrolysis only when endo-xylanase activity was followed by that of xylanase A. |
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