SUMMARY: The deacetoxycephalosporin C (DAOC) synthase (expandase) of was highly purified, as shown by SDS-PAGE and isoelectric focusing. The enzyme catalysed the oxidative ring expansion that converts penicillin N into DAOC. The enzyme was very unstable but could be partially stabilized in 25 m-Tris/HCl, pH 9·0, in the presence of DTT (0·1 m). The enzyme required 2-oxoglutarate, oxygen and Fe, but did not need ATP, ascorbic acid, Mg or K. The optimum temperature was between 25 and 30 °C. The DAOC synthase showed a high specificity for the penicillin substrate. Only penicillin N but not isopenicillin N, penicillin G or 6-aminopenicillanic acid served as substrates. 2-Oxoglutarate analogues were not used as substrates although 2-oxobutyrate and 3-oxoadipate inhibited the enzyme by 100% and 56% respectively. The enzyme was strongly inhibited by Cu, Coand Zn. The apparent values for penicillin N, 2-oxoglutarate and Fewere 52 μ, 3 μand 71 μrespectively. The enzyme was a monomer with a molecular mass of 27000 Da ± 1000.


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