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 gene identified and further defines a new subfamily within the flavoprotein disulfide oxidoreductases. The three STHs identified all lack one of the redox 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 fluorescens and Escherichia coli STH. Chimeric STHs were constructed which showed that the C-terminal legion is important for polymer formation. The ii. vinelandii 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 Societies, Published by Elsevier Science B.V. Ail rights reserved.