ROLE OF SCAFFOLDING PROTEIN CIPC OF CLOSTRIDIUM-CELLULOLYTICUM IN CELLULOSE DEGRADATION

The role of a miniscaffolding protein, miniCipC(1), forming part of Cl ostridium cellulolyticum scaffolding protein CipC in insoluble cellulo se degradation was investigated, The parameters of the binding of mini CipC(1), which contains a family III cellulose-binding domain (CBD), a hydrophilic domain, and a cohesin domain, to four insoluble cellulose s were determined, At saturating concentrations, about 8.2 mu mol of p rotein was bound per g of bacterial microcrystalline cellulose, while Avicel, colloidal Avicel, and phosphoric acid-swollen cellulose bound 0.25, 0,38, and 0.55 mu mol of miniCipC(1) per g, respectively. The di ssociation constants measured varied between 1.3 x 10(-7) and 1.5 x 10 (-8) M. These results are discussed with regard to the properties of t he various substrates. The synergistic action of miniCipC(1) and two f orms of endoglucanase CelA (with and without the dockerin domain [CelA (2) and CelA(3), respectively]) in cellulose degradation was also stud ied. Although only CelA, interacted with miniCipC(1) (K-d, 7 x 10(-9) M), nonhydrolytic miniCipC(1) enhanced the activities of endoglucanase s CelA(2) and CelA(3) with all of the insoluble substrates tested, Thi s finding shows that miniCipC(1) plays two roles: it increases the enz yme concentration on the cellulose surface and enhances tile accessibi lity of the enzyme to the substrate by modifying the structure of the cellulose, leading to an increased available cellulose surface area. I n addition, the data obtained with a hybrid protein, CelA(3)-CBDCipC, which was more active towards all of the insoluble substrates tested c onfirm that the CBD of the scaffolding protein plays an essential role in cellulose degradation.