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Cellulosic microfibrils in plant cell walls are largely ensheathed and probably tethered by hydrogen-bonded hemicelluloses. Ensheathing may vary developmentally as hemicelluloses are peeled to enable cell expansion. We characterised a simple method to quantify ensheathed versus naked cellulosic surfaces based on the ability to adsorb a radiolabelled ‘cellulose-complementary oligosaccharide’, [³H]cellopentaitol. Filter-paper (cellulose) adsorbed 40% and >80% of aqueous 5 nM [³H]cellopentaitol within ∼1 and ∼20 h respectively. When [³H]cellopentaitol was rapidly dried onto filter-paper, ∼50% of it was desorbable by water, whereas after ∼1 day annealing in aqueous medium the adsorption became too strong to be reversible in water. ‘Strongly’ adsorbed [³H]cellopentaitol was, however, ∼98% desorbed by 6 M NaOH, ∼50% by 0.2 M cellobiose, and ∼30% by 8 M urea, indicating a role for hydrogen-bonding reinforced by complementarity of shape. Gradual adsorption was promoted by kosmotropes (1.4 M Na2SO4 or 30% methanol), and inhibited by chaotropes (8 M urea), supporting a role for hydrogen-bonding. [³H]Cellopentaitol adsorption was strongly competed by non-radioactive cello-oligosaccharides (Cell2–6), the IC50 (half-inhibitory concentration) being highly size-dependent: Cell2, ∼70 mM; Cell3, ∼7 mM; and Cell4–6, ∼0.05 mM. Malto-oligosaccharides (400 mM) had no effect, confirming the role of complementarity. The quantity of adsorbed [³H]cellopentaitol was proportional to mass of cellulose. Of seven cottons tested, wild-type Gossypium arboreum fibres were least capable of adsorbing [³H]cellopentaitol, indicating ensheathment of their microfibrillar surfaces, confirmed by their resistance to cellulase digestion, and potentially attributable to a high glucuronoarabinoxylan content. In conclusion, [³H]cellopentaitol adsorption is a simple, sensitive and quantitative way of titrating ‘naked’ cellulose surfaces. [ABSTRACT FROM AUTHOR]

Separate binding of several purified cellulolytic components of Trichoderma reesei on to filter paper was studied and concomitant hydrolysis rates evaluated. Enhancement of mass transfer from the bulk liquid to the solid substrate by agitation has two different effects on adsorption depending on the type of enzyme: (i) the fraction of cellobiohydrolase II (CBH II) and endoglucanase III (EG III) bound at equilibrium is increased, whereas (ii) the rate but not the extent of cellobiohydrolase I (CBH I) and endoglucanase I (EG I) adsorption is affected. The adsorption of CBH I core, a component lacking the cellulose-binding domain (CBD), is, however, not significantly influenced by mass transfer. The CBH I interdomain peptide (present in CBH I core b) does not participate in adsorption but enhances stability. The adsorption of CBH I core proteins is a fully reversible process whereas that of the intact CBH I is not. Thus, the interaction of the CBD with filter paper apparently accounts for the mass-transfer-limited binding rate and also for the irreversible adsorption of intact CBH I. Adsorption isotherms at 50 degrees C indicate very similar relative association constants for the intact cellulases (0.24-0.30 l/g of cellulose), but drastically reduced values for CBH I core proteins (0.03 l/g of cellulose). The specific activities of adsorbed CBH I and of its core proteins are identical and a linear relationship between adsorption and rates of hydrolysis is found only for these enzymes. Thus, non-productive binding on to cellulose seems evident in the case of CBH II and EG III but not CBH I.

The relationship between the electrophoretic mobility, the molecular weight and the charge of peptides at pH1.9 was re-investigated. It was shown that the relationship at pH1.9 is quantitatively similar to that which holds at pH6.5.