A small number of pectin binding proteins were obtained by passing soluble proteins from Cucurbita pepo L. hypocotyls through a column containing polygalacturonic acid (PGA) entrapped within polyacrylamide gel. These proteins were bound significantly to the gel only in the presence of Ca²⁺ and could be eluted with NaCL Two of these proteins were apparently immunologically related to human vitronectin. A comparison with heparinagarose showed that the animal glycosaminoglycan retained the same proteins than Ca²⁺ -PGA/polyacrylamide. In order to understand how these proteins interacted with the anionic polysaccharides, the binding to Ca²⁺ -PGA of anionic and cationic isoperoxidases (EC 1.11.1.7), belonging to the set of pectin binding proteins, was measured in the presence of various additions. For this purpose, we used a binding test based on the co-sedimentation of peroxidases and Ca²⁺ -PGA gel upon centrifugation. The linkage between peroxidases and Ca²⁺ -PGA was broken by increasing the ionic strength with NaCl or by adding various cationic polyamino acids. To be inhibitory, polycations must be long enough and have a defined conformation. Chitosan was ineffective. Active polycations did not break the Ca²⁺ -induced polymerization of PGA, but competed with the proteins for the binding to PGA molecules in their Ca²⁺ -mediated conformation. Experiments with the protein modifiers sulfo-NHS-acetate and phenylglyoxal showed that the amino acids lysine and arginine were involved in the binding mechanism, even in the case of the anionic isoperoxidase. It may be envisaged that this binding mechanism could allow some extracellular enzymes or proteins to be properly distributed within the extracellular matrix, in order to exert their catalytic functions on selected sites.