ISOLATION AND PRELIMINARY CHARACTERIZATION OF THE SUBUNITS OF THE TERMINAL COMPONENT OF NAPHTHALENE DIOXYGENASE FROM PSEUDOMONAS-PUTIDA NCIB 9816-4

The terminal oxygenase component (ISP(NAP)) of naphthalene dioxygenase from Pseudomonas putida NCIB 9816-4 was purified to homogeneity. The protein contained approximately 4 g-atoms each of iron and acid-labile sulfide per mol of ISP(NAP), and enzyme activity was stimulated signi ficantly by addition of exogenous iron. The large (alpha) and small (b eta) subunits of ISP(NAP) were isolated by two different procedures. T he NH2-terminal amino acid sequences of the alpha and beta subunits we re identical to the deduced amino acid sequences reported for the ndoB and ndoC genes from P. putida NCIB 9816 and almost identical to the N H2-terminal amino acid sequences determined for the large and small su bunits of ISP(NAP) from P. putida G7. Gel filtration in the presence o f 6 M urea gave an alpha subunit with an absorption maximum at 325 nm and broad absorption between 420 and 450 nm. The alpha subunit contain ed approximately 2 g-atoms each of iron and acid-labile sulfide per mo l of the subunit. The beta subunit did not contain iron or acid-labile sulfide. These results, taken in conjunction with the deduced amino a cid sequences of the large subunits from several iron-sulfur oxygenase s, indicate that each alpha subunit of ISP(NAP) contains a Rieske [2Fe -2S] center.