CHANGES IN THE MOLECULAR-SIZE DISTRIBUTION OF INSOLUBLE CELLULOSES BYTHE ACTION OF RECOMBINANT CELLULOMONAS-FIMI CELLULASES

Specific patterns of attacks of cotton, bacterial cellulose and bacter ial microcrystalline cellulose (BMCC) by recombinant cellulases of Cel lulomonas fumi were investigated. Molecular-size distributions of the celluloses were determined by high-performance size-exclusion chromato graphy. Chromatography of cotton and bacterial celluloses revealed sin gle major peaks centred over progressively lower molecular-mass positi ons during attack by endoglucanase CenA. In advanced stages, a second peak appeared at very low average size (approx. 11 glucosyl units); ul timate weight losses were approximately 30%. The isolated catalytic do main of CenA, p30, gave results very similar to those with complete Ce nA. CenA did not effectively depolymerize or solubilize BMCC significa ntly. Molecular-size distributions of cotton and bacterial cellulose i ncubated with endoglucanases CenB or CenD exhibited one major peak reg ardless of incubation time; low-molecular-mass fragments did not accum ulate. Weight losses were 40 and 35% respectively. The single peak shi fted to lower-molecular-mass positions as incubation continued, but hi gh-molecular-mass material persisted. CenB and CenD readily attacked a nd solubilized BMCC (approx. 70%). We conclude that CenA attacks cellu lose by preferentially cleaving completely through the cellulose micro fibrils at the amorphous sites, and much more slowly by degrading the crystalline surfaces. Conversely, CenB and CenD cleave the amorphous r egions much less efficiently while vigorously degrading the surfaces o f the crystalline regions of the microfibrils.