Interaction of Leptospira interrogans ferredoxin-NADP(H) reductase with ferredoxin

ROSSI, MA. AGUSTINA; LÓPEZ RIVERO, ARLETH; CECCARELLI, EDUARDO A; CATALANO DUPUY, DANIELA L

Rosario

Congreso; L Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2014

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Leptospira interrogans is a parasitic bacterium that infects humans and causes leptospirosis. L. interrogans contains a plastidic-typeferredoxin-NADP+ reductase (LepFNR) similar to the enzymes presentin plant plastids and different from the bacterial-type FNRs. A unique feature foundin LepFNR structure is a loop that ranges from I73 to R94. By structuralmodeling we noticed that this structure may interfere with the ferredoxin (Fd)binding. Also, a conserved lysine essential for productive binding of Fd toplant FNRs, corresponds to a threonine residue (T92) in LepFNR. This allow usto suggest that the binding of Fd LB107, identified as a substrate of LepFNRshould take place by a different arrangement from the observed in plantenzymes. We designed and constructed different site-directed mutants of LepFNRaltering the subdomain I73-R94. We succeeded in expressing and purifying themutants proteins properly folded and with the FAD bound. We measured thediaphorase activity catalyzed by the LepFNR variants. We found that the substitutionsdid not lead to mayor changes in this activity and in the NADP(H) binding. Wewere also able to measure Fd-dependant cytochrome c reductase activity with the mutants. Our results indicate thatthe LepFNR subdomain I73-R94 participates in Fd binding and intervenes insubstrate specificity.