Streptomyces strains as a source of enzymes, able to hydrolyse the Clavibacter exopolymer, clavan

Two soil isolates, identified as Streptomyces sp. and designated as Streptomyces sp. YSDL-10 and Streptomyces sp. YSDL-20, were among the few microorganisms and able to hydrolyse and grow on clavan as sole carbon source. This biopolymer, very rich in L-fucose, is produced in by Clavibacter michiganensis subsp. michiganensis LMG 5604. The enzyme activity tentatively called clavanase was mainly detected in the culture liquid. However, qualitative tests showed that this activity was very weak in the supernatant. In order to test whether this activity was dependent of induction, several homo- and heteropolysaccharides as well as clavan were used as sole carbon source: the strains were grown on 10 g/l clavan, starch, cellulose, pectin, polygalacturonic acid, xylan or starch. After 5 days of growth, samples were centrifuged (10.000 rpm, 20 min) and the supernatants were lyophilised This lyophilised preparation was used as crude enzyme source. This preparation was dissolved in 500 microl phosphate buffer (pH 7.0) and added to 500 microl of clavan (0.5% w/v). Upon incubation of this mixture at 30 degrees C during 30 minutes, reducing sugars and L-fucose levels were quantified. The clavanase activity was relatively high in terms of reducing sugar content, but was weak in terms of L-fucose liberated. Nevertheless, qualitative tests performed on solid screening clavan medium showed large clearing zones around Streptomyces colonies. This indicates that clavan is split into oligosaccharides with sugar reducing moieties. The glycosidase activities detected in Czapek Dox-clavan medium were also weak, except for beta-D-xylosidase. This suggests that this enzyme activity could have some role in the clavanase enzyme complex.