INDUCTION, LOCALIZATION, AND PURIFICATION OF A NOVEL SIALIDASE, DEAMINONEURAMINIDASE (KDNASE), FROM SPHINGOBACTERIUM-MULTIVORUM

Recently, we reported the discovery of a new type of sialidase, KDNase , which specifically hydrolyzes the ketosidic linkages of 2-keto-3-deo xy-D-glycero-D-galactonononic acid (KDN), but not N-acylneuraminyl lin kages. We now report that this enzyme, designated KDNase SM, is an ind ucible enzyme that is localized in the periplasm of Sphingobacterium m ultivorum. Growth of S. multivorum in the presence of KDN-containing o ligosaccharide alditols, KDN alpha 2-->3Gal beta 1-->3GalNAc alpha 1-- >3[KDN alpha 2-->(8KDN alpha 2-->)(n)-->6]GalNAcol, as a sole carbon s ource induced KDNase SM activity 15-40-fold, compared with growth in t he absence of inducer, KDN, Neu5Ac, or Neu5Ac oligomers were ineffecti ve as inducers. The enzyme was released from the periplasm of induced cells by cold osmotic shock and purified 700-fold to homogeneity. The specific activity of the pure enzyme was 82,100 units/mg of protein, K DNase SM activity resided in a single polypeptide chain with an estima ted molecular weight of approximately 47,500. Enzyme activity was maxi mal at near neutral pH. The availability of pure KDNase will now make it possible to study the structure and functional role of KDN-glycocon jugates and to determine the molecular mechanism whereby the enzyme ca n discriminate between KDN and N-acylneuraminic acid.