The importance of crystal lattice structure of cellulose in the production and action of cellulase

Abstract

The production and action of Trichoderma viride enzymes capable of degrading crystalline forms of cellulose were studied. The four stable crystal forms (I, II, IV and II/IV) were used in the growth medium, both before and after the removal of the amorphous material. Various enzyme preparations capable of solubilizing crystalline cellulose forms were thus obtained. Only crystalline forms were used as substrates. Different experimental approaches were taken to determine the effects of crystal structure on the induction and action of "cellulase.” The study was also aimed to obtain evidence from these experiments which may be helpful in elucidating the mechanism of the degradation of crystalline cellulose. The results of various experiments clearly demonstrated the importance of the crystal structure. Thus a difference in growth and the protein production was observed when the growth medium contained different crystal forms of cellulose. The measurements of the specific activities of various enzyme-substrates revealed differences relative to the crystal forms. The thermal inactivation of various enzyme preparations gave different rates of inactivation. The rates of inactivation were also dependent on the type of crystalline substrate used in the assay. The measurement of the activation energies of various enzyme-substrate combinations indicated that the enzyme produced in the presence of a given substrate was also most suitable to degrade that particular form. This showed that different enzymes or enzyme systems were induced by different crystal forms. The activation energy measurements also indicated that different crystal forms were energetically unequal. The crystal form I, a natural modification, had the lowest energy. The various crystal forms thus differed only relatively, all of them still following the same basic reaction pattern. This was demonstrated by x-ray studies which showed that the crystal structure as well as the crystallinity of all the forms remained unchanged during the course of reaction. Electron microscopic studies of the degraded and undegraded crystalline cellulose I indicated that the enzymatic action involved fragmentation of the substrate initiated by the formation of fissures parallel to the long axes of crystallite aggregates. The results obtained by using the substrates containing particles of different sizes, supported the observation (40) that there exists a direct proportionality between substrate surface area and the rate of enzymic solubilization of crystalline cellulose. These results also supported the fragmentation phenomenon indirectly. A mechanism for the degradation of crystalline cellulose is proposed.