Cloning of the sth gene from Azotobacter vinelandii and construction of chimeric soluble pyridine nucleotide transhydrogenases

The gene encoding the soluble pyridine nucleotide transhydrogenase (STH) of Azotobacter vinelandii was cloned and sequenced. This is the third sth gen e identified and further defines a new subfamily within the flavoprotein di sulfide oxidoreductases. The three STHs identified all lack one of the redo x active cysteines that are characteristic for this large family of enzymes , and instead they contain a conserved threonine residue at this position. The recombinant A. vinelandii enzyme was purified to homogeneity and shown to form Filamentous structures different from those of Pseudomonas fluoresc ens and Escherichia coli STH. Chimeric STHs were constructed which showed t hat the C-terminal legion is important for polymer formation. The ii. vinel andii STH containing the C-terminal region of P. fluorescens or E. coli STH showed structures resembling those of the STH contributing the C-terminal portion of the protein. (C) 2000 Federation of European Microbiological Soc ieties, Published by Elsevier Science B.V. Ail rights reserved.