Non-specific protein binding by adsorbents designed for the specific affinity chromatography of sialidase in crude bacterial extracts.

A number of derivatives of Sepharose 4B gels were prepared containing tyramine, -Gly -Gly -Tyr and the tetrapeptide, Thr (But)-Phe-Pro-Tyr -. A specific inhibitor of sialidase, p-diazo-phenyl oxamic acid, was also coupled to these phenolsubstituted Sepharose gels. The effectiveness of these gels to purify sialidase by "affinity" chromatography was examined. The sialidase from Vibrio cholerae was adsorbed by these gels and subsequently re-eluted by a change in pH and temperature. The adsorption and elution of enzyme was independent of the presence or absence of the "specific inhibitor" of the sialidase, phenyl oxamic acid. A mechanism for the adsorption and subsequent elution of the sialidase is proposed. It is important to re-evaluate the biological data obtained with sialidase prepared by the affinity column recommended by Cuatrecasas and Illiano, Biochem. Biophys. Res. Commun. 44, 178-184 (1971), since glycosidases from Clostridium perfringens other than sialidase are also adsorbed and eluted under the conditions specified.