The Ferredoxin NADP+ Reductase from Escherichia coli contains NADP+ tightly bound

MATÍAS A. MUSUMECI; DANIELA L. CATALANO-DUPUY; EDUARDO A. CECCARELLI

Flavins and Flavoproteins 2011

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Lugar: New York; Año: 2012; p. 50 - 55

Ferredoxin-NADP+ reductases (FNRs, EC 1.18.1.2) are ubiquitous enzymes which function as switch in redox reactions between NADPH and ferredoxin or flavodoxin. This feature of catalyzing reactions involving one and two electron carriers is possible thanks to their FAD prosthetic group, which can exist in different redox states. In plants FNRs have evoluted to produce high amounts of NADPH during photosynthesis. However, in bacteria they favour the inverse reaction, namely the reduction of ferredoxin or flavodoxin from NADPH. Ih this work we have performed a comparative analysis of the NADP+ binding mode in pea FNR (a plastidic type FNR) and E. coli FNR (a bacterial type FNR). Through HPLC analysis, we have observed that a fraction of molecules of the population of E. coli FNR are able to bind NADP+ when are expressed in E. coli cells and retain it tightly during the purification procedures, at difference of pea FNR. The bacterial FNR containing NADP+ could be separated from the FNR without nucleotide and differences in the spectra were observed. Studies of NADP+ binding in the time suggeted that the NADP+ binding mode could change slowly. In order to obtain more information the interaction of these FNRs with P-AMP was also analyzed. Thus, by comparing the obtained results for NADP+ and P-AMP interaction we found that the nicotinamide portion contribute to the binding of the NADP+ in E. coli FNR but it is not the case in pea FNR. The implication of this particular NADP+ binding mode in E. coli FNR in the phisiological function of the enzyme is discussed.