Oxidative scission of plant cell wall polysaccharides by ascorbate-induced hydroxyl radicals

Abstract

Scission of plant cell wall polysaccharides in vivo has generally been assumed to be enzymic. However, in the presence of l-ascorbate, such polysaccharides are shown to undergo non-enzymic scission under physiologically relevant conditions. Scission of xyloglucan by 1 mM ascorbate had a pH optimum of 4.5, and the maximum scission rate was reached after a 10–25-min delay. Catalase prevented the scission, whereas added H₂O₂ (0.1–10 mM) increased the scission rate and shortened the delay. Ascorbate caused detectable xyloglucan scission above approx. 5 µM. Dehydroascorbate was much less effective. Added Cu²⁺ (> 0.3 µM) also increased the rate of ascorbate-induced scission; EDTA was inhibitory. The rate of scission in the absence of added metals appeared to be attributable to the traces of Cu (2.8 mg·kg^-1) present in the xyloglucan. Ascorbate-induced scission of xyloglucan was inhibited by radical scavengers; their effectiveness was proportional to their rate constants for reaction with hydroxyl radicals (^•OH). It is proposed that ascorbate non-enzymically reduces O₂ to H₂O₂, and Cu²⁺ to Cu⁺, and that H₂O₂ and Cu⁺ react to form ^•OH, which causes oxidative scission of polysaccharide chains. Evidence is reviewed to suggest that, in the wall of a living plant cell, Cu⁺ and H₂O₂ are formed by reactions involving ascorbate and its products, dehydroascorbate and oxalate. Systems may thus be in place to produce apoplastic ^•OH radicals in vivo. Although ^•OH radicals are often regarded as detrimental, they are so short-lived that they could act as site-specific oxidants targeted to play a useful role in loosening the cell wall, e.g. during cell expansion, fruit ripening and organ abscission.