2xnc

Publication Abstract from PubMed

Plant-type ferredoxin-NADP(H) reductases (FNRs) group in two classes, plastidic with an extended FAD conformation and high catalytic rates, and bacterial with a folded flavin nucleotide and low turnover rates. The 112-123 beta-hairpin from a plastidic FNR and the carboxy-terminal tryptophan of a bacterial FNR, suggested to be responsible for the FAD differential conformation, were mutually exchanged. The plastidic FNR lacking the beta-hairpin was unable to fold properly. An extra tryptophan at the carboxy terminus, emulating the bacterial FNR, resulted in an enzyme with decreased affinity for FAD and reduced diaphorase and ferredoxin-dependent cytochrome c reductase activities. The insertion of the beta-hairpin into the corresponding position of the bacterial FNR increased FAD affinity although not affecting its catalytic properties. The same insertion with simultaneous deletion of the carboxy-terminal tryptophan produced a bacterial chimera emulating the plastidic architecture with increased <i>k</i>_cat and catalytic efficiency for the diaphorase activity and a decrease in the enzyme's capability to react with its substrates ferredoxin and flavodoxin. Crystallographic structures of the chimeras showed no significant changes in their overall structure, although alterations in the FAD conformations were observed. Plastidic and bacterial FNRs thus reveal differential effects of key structural elements. While the 112-123 beta-hairpin modulates catalytic efficiency of plastidic FNR it seems not to affect the bacterial FNR behaviour, which instead can be improved by the loss of the C-terminal tryptophan. This report highlights the role of the FAD moiety conformation and the structural determinants involved in stabilizing it, ultimately modulating the functional output of FNRs.

Swapping FAD Binding Motifs Between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases.,Musumeci MA, Botti H, Buschiazzo A, Ceccarelli EA Biochemistry. 2011 Feb 9. PMID:21306142^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.