The coenzyme binding site of bacterial Ferredoxin/flavodoxin NADP(H) reductases.

INMACULADA PEREZ-DORADO; JUAN. A HERMOSO; GUILLERMINA GOÑI; MILAGROS MEDINA; ANA BORTOLOTTI; NÉSTOR CARRILLO; NÉSTOR CORTEZ

Jaca, Spain

Simposio; 16th Symposiumon Flavins and Flavoproteins; 2008

Flavoproteins with ferredoxin-NADP(H) reductase (FPR/FNR) activity are ubiquitous among living organisms, participating in electron transport pathways that require splitting of electrons between obligatory two-electron carriers (i. e., pyridine nucleotides) and one-electron transporters such as ferredoxin or flavodoxin [1]. They are found in both strict and facultative anaerobes, suggesting an ancient origin. After appearance of oxygenic photosynthesis, FPRs were recruited into the photosynthetic electron transport chain, direction of electron flow was reserved to favour NADP+ reduction, and an increase in catalytic efficiency occurred to cope with the demands of the novel metabolism [1]. Enzymes found in plastids and cyanobacteria are characterized by an extended FAD conformation and an invariant tyrosine residue at the C-terminus. Instead, the FPRs from eubacteria bind FAD in a bent conformation, and the amino acid facing the flavin may be aromatic or aliphatic. To acknowledge the distinctive features of the two reductase classes, the name FNR has been to the plastidic/cyanobacterial enzymes, while those found in eubacteria are termed FPRs [1]. A slower hydride transfer rate and a weaker coenzyme binding were previously described for FPRs when compared with their plastidic counterparts [1]. Structurally-based aligments and model docking described herein indicate that improvement of NADP(H) interaction was attained through stronger nucleotide binding, whereas single turnover experiments showed that the general reaction pathway was hardly modified. Contacts with the adenosine moiety of NADP(H) play a key role in docking. The most remarkable difference between the two types of reductases concerns R203 of Rhodobacter FPR, which is replaced by a conserved tyrosine in FNRs. This tyrosine appears to be critical for nucleotide affinity undergone by alanine mutants in Anabaena FNR [2]. Replacement of R203 by a tyrosine would turn a possible cation-π  interaction into a π- π interaction with the adenosine, and it is likely that this substitution was one of the key steps leading to increased catalytic competence in photosynthetic FNRs.