Acidified sodium chlorite cleaves isodityrosine and solubilizes covalently bound hydroxyproline-rich material from cell walls. This has been taken as evidence that isodityrosine acts as a cross-link holding the hydroxyproline-rich glycoprotein extensin in the cell wall. However, acidified chlorite was found to cleave peptide bonds in salt-soluble extensin and in bovine serum albumin (BSA). This invalidates the use of conventional acidified chlorite treatment to provide evidence for isodityrosine crosslinks. The rafto of BSA:chlorite was important in determining peptidyl cleavage. At a ratio of 0.75:1.00 (mole amino acid residues/mole chlorite), or higher, peptidyl cleavage was not detected. Furthermore, in samples where a low concentration of radioactive extensin was present, BSA substantially protected the peptide bonds of the extensin against peptidyl cleavage during treatment with acidified chlorite, while not preventing the cleavage of isodityrosine. Therefore, acidified sodium chlorite plus BSA was a more specific reagent for the cleavage of isodityrosine than was acidified chlorite alone. This modified treatment solubilized in intact form the 'covalently bound' extensin from cell walls of Capsicum frutescens (chili pepper) suspension cultures, providing new evidence compatible with the view that extensin molecules are held in the cell wall by isodityrosine cross-links.Plant cell walls contain the HRGP2 called extensin, which forms an insoluble and perhaps structurally important network (13). Mature extensin is insoluble in many of the usual protein extractants such as detergents (8), salts (24), dilute acids and alkalis (14), PAW (present paper), and anhydrous hydrogen fluoride (16) and is therefore thought to be covalently bound within the cell wall. This insolubility hindered the study ofextensin until the isolation ofsalt-soluble extensin precursors (monomers ofMr about 9 x 104) from cell suspension cultures (21) and wounded carrot disks (4, 23), which has enabled progress on extensin's structure and properties to be made in recent years (see review, ref. 26 the wall network is unknown, but the isolation ofthe phenolic amino acid isodityrosine (an oxidatively coupled dimer of tyrosine units linked by a diphenylether bridge) from cell wall hydrolysates (8) led to the suggestion that isodityrosine could form inter-polypeptide cross-links between extensin molecules (2, 9).Studies on isolated carrot root disks (4,5) showed that newly synthesized extensin monomers were secreted from the cytoplasm and were slowly insolubilized in the walls. Insolubilization was accompanied by isodityrosine formation, but it was not proven that the isodityrosine formed was responsible for insolubilization.In another study HRGP was solubilized from bean cell walls by treatment with acidified chlorite (17,19,20), a reagent that is extensively used for delignification and known to split phenolic linkages (6). In addition to an ability to solubilize HRGP, acidified chlorite has been shown to degrade pure isodityrosine (8,...