Nonidentical subunits of protocatechuate 3,4-dioxygenase

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Abstract
Protocatechuate 3,4-dioxygenase (EC 1.13.11.3) [of Pseudomonas aeruginosa] was reported to have a MW of 700,000 and to consist of 8 identical subunits, each containing 1 atom of Fe2+ and a substrate binding site. This subunit dissociates further into 4 smaller subunits of 2 nonidentical types (.alpha.2.beta.2), upon sodium dodecyl sulfate gel electrophoresis. The MW of the .alpha. and .beta. subunits were estimated to be 22,500 and 25,000, respectively. Isoelectric focusing of the enzyme in 6 M urea revealed that the isoelectric points of the .alpha. and .beta. subunits were 5.2 and 9.5, respectively. Separation of the 2 subunits was achieved by chromatography on sulfopropyl (SP)-Sephadex in 6 M urea after treatment of the enzyme with 8 M urea at 37.degree. C for 6 h. The NH2-terminal sequence of the .alpha. subunit was determined to be Pro-Ile-Glu-Leu-Leu-Pro-Glu-Thr-Pro-Ser-Glx-Thr-Ala-Gly and that of the .beta. subunit, Pro-Ala-Gln-Asp-Asn-Ala-Arg-Phe-Val-Ile-Arg-Asx-Arg-Asx. Phenylalanine was the COOH-terminal residue of the .alpha. subunit. The COOH terminus of the .beta. subunit was not detected by any of 3 methods employed.