Hexopyranoside: Cytochrome c Oxidoreductase from Agrobacterium tumefaciens

Abstract

An inducible enzyme, catalyzing the formation of 3‐ketosugars, has been isolated in a pure state from Agrobacterium tumefaciens, grown on lactose. It occurs in the cytoplasm. It is a FAD‐protein with a sedimentation coefficient of 5.1 S and a molecular weight of 85,000 × 7,700. The pH optimum varies with the nature of the electron acceptor: with cytochrome c it is 7.0, with 2,6‐dichlorophenolindophenol 6.0 and with 5‐methylphenazinium methyl sulphate 7.9. p‐Chloromercuribenzoate, and to some extent atabrine, have an inhibitory effect. In view of its lability, the enzyme is best preserved in a nitrogen atmosphere at pH 7 and 0°. The substrate specificity as studied with a variety of sugars, justifies the name of hexopyranoside: cytochrome c oxidoreductase for the enzyme. As a rule, disaccharides are more easily oxidized than monosaccharides. The highest affinity is observed with cellobiose and lactobionic acid (Km=2.0 × 10‐4M). Most of the 3‐ulose endproducts were detected by thin layer and/or polarography. The minimal structural requirements of a substrate sugar allow the proposal of a molecular model for the active centre of the enzyme. The oxidation‐reduction potential at pH 7 and 25° of the system lactose/3‐ketolactose is calculated to be Eo=+0.16 V. Copyright © 1968, Wiley Blackwell. All rights reserved