| Abstrakt: |
Two membrane-bound nitrate reductases, NRA and NRZ, exist in Escherichia coli. Both isoenzymes are composed of three structural subunits, α, β and γ encoded by narG/narZ, narH/narY and narl/narV, respectively. The genes are in transcription units which also contain a fourth gene encoding a poiypeptide, δ, which is not part of the final enzyme. A strain which is devoid of, or does not express, the nar genes, was used to investigate the role of the δ and γ polypeptides in the formation and/or processing of the nitrate reductase. When only the α and β polypeptides are produced, an (αβ) complex exists which is inactive and soluble. When the α, β and δ polypeptides are produced, the (αβ) complex is active with artificial donors such as benzyl viologen but is soluble. When the α, β and γ polypeptides are produced, the (αβ) complex is inactive but partially binds the membrane. It was concluded that the y polypeptide is involved in the binding of the (αβ) complex to the membrane while the 6 polypeptide is indispensable for the (αβ) nitrate reductase activity. The activation by the 5 polypeptide does not seem to involve the insertion of the redox centres of the enzyme since the purified inactive (αβ) complex was shown to contain the four iron-sulphur centres and the molybdenum cofactor, which are normally present in the native purified enzyme. The extreme sensitivity of this inactive complex to thermal denaturation or tryptic treatment favours the idea that the 5 potypeptide promotes the correct assembly of the a and p subunits. Although this corresponds to the definition of a chaperone protein this possibility has been rejected. In this study we have also demonstrated that the 3 or 7 polypeptide encoded by one nar operon can be substituted succesfully for by its respective counterpart from... [ABSTRACT FROM AUTHOR] |
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