[Degradation of lignin-carbohydrate substrate by soil fungi--producers of laccase and cellobiose dehydrogenase]. / Razlozhenie lignouglevodnogo substrata pochvennymi gribami--produtsentami lakkazy i tsellobiozodegidrogenazy.

The growth of nonsporulating mycelial fungi INBI 2-26(+), producer of laccase; INBI 2-26(-), producer of cellobiose dehydrogenase; and their mixed culture on lignin-carbohydrate substrates under conditions of submerged fermentation were studied. The degrees of degradation of lignin, cellulose, and hemicellulose of cut straw over 23 days amounted to 29.8, 51.4, and 72% for the laccase producer; 15.8, 33.9, and 59.1% for the cellobiose dehydrogenase producer; and 15.8, 39.4, and 64.5% for the mixed culture, respectively. The laccase activity in the medium when strain 2-26(+) was cultivated individually reached its maximum on day 28; the activity of cellobiose dehydrogenase of strain 2-26(-), on days 14 to 28. A method for determining cellobiose dehydrogenase activity in the presence of laccase was developed. In the mixed culture, both enzymes were formed; however, the level of laccase synthesis was 1.5-fold lower compared to that of strain 2-26(+), while synthesis of cellobiose dehydrogenase was similar to that of the corresponding producer. Cellobiose dehydrogenase failed to boost the action of laccase while degrading the lignin of straw.

[Isolation and characterization of a cellobiose dehydrogenase formed by a asporogenic mycelial fungus INBI 2-26(-)]. / Vydelenie i kharakteristika tsellobiozodegidrogenazy, obrazuemoi nesporuliruiushchim mitselial'nym gribom INBI 2-26(-).

A nonsporulating fungus isolated from dioxine-containing tropical soils forms cellobiose dehydrogenase, when grown in media supplemented by a source of cellulose. The enzyme purified to homogeneity by SDS-PAGE (yield, 43%) had an M(r) of 95 kDa; its pH optimum was in the range 5.5-7.0; more than 50% activity was retained at pH 4.0-8.0 (citrate-phosphate buffer). The absorption spectrum of the enzyme in the visible range had the characteristic appearance of flavocytochrome proteins. Cellobiose dehydrogenase oxidized cellobiose and lactose (the respective K(M) values at pH 6.0 equaled 4.5 +/- 1.5 and 56 microM) in the presence of dichlorophenolindophenol (K(M) app = 15 +/- 3 microM at pH 6.0) taken as an electron acceptor. Other sugars were barely if at all oxidized by the enzyme. Neither ethyl-beta-D-cellobioside, heptobiose, nor chitotriose inhibited the enzymatic oxidation of lactose, even under the conditions of 100-fold molar excess. The enzyme was weakly inhibited by sodium azide dichlorophenolindophenol reduction and exhibited affinity to amorphous cellulose. At 55 degrees C and pH 6.0 (optimum stability), time to half-maximum inactivation equaled 99 min. The enzyme reduced by cellobiose was more stable than the nonreduced form. Conversely, the presence of an oxidizer (dichlorophenolindophenol) decreased the stability eight times at pH 6.0. In addition, the enzyme acted as a potent reducer of the single-electron acceptor cytochrome c3+ (K(M) app = 15 microM at pH 6.0).