C-terminus mutation effects on spectroscopic properties of Rhodobacter capsulatus ferredoxin/flavodoxin NADP reductase

M. L. VALLE; I. ABATEDAGA; F. E. MORÁN VIEYRA; C.D. BORSARELLI; N. CORTEZ

San Javier, Tucumán

Congreso; XLI REUNION ANUAL DE LA SOCIEDAD ARGENTINA DE BIOFISICA; 2012

Sociedad Argentina de Biofísica

The photosynthetic bacterium Rhodobacter capsulatus contains a single (flavodoxin)-NADP(H) oxidoreductase (RcFPR) that catalyzes electron transfer from NADP(H) to the flavodoxin nifF for nitrogenase reduction. The X-ray crystallography of the enzyme showed two domains harboring the FAD and NADP(H). Plastidic class of reductases presents a C-terminal Tyr residue involved in catalytic mechanism and stabilizing the RE face of the isoalloxazine ring of FAD. Differently, bacterial class FPRs carry a Phe or Ala instead terminal Tyr, and a C-terminal extension -FVGEGI, which also participates in NADP(H) binding and hydride transfer mechanisms (1). A site-directed mutagenesis approach on RcFPR was performed, producing three different mutants: A266Y, A266∆ and A266Y∆, which modifies both structural and catalytic properties . In this work,the UV-Vis absorption and fluorescence properties of FAD in the mutant series was analyzed. The results indicate the role of C-terminal extension -FVGEGI in the modulation of FAD exposure and stability in RcFPR. Fluorescence parameters: quantum yield (ΦF), steady state anisotropy (), lifetimes (τi), lifetime percentage contribution (αi)