Pulse radiolysis and related studies on the Ru-modified His 16 derivative of Anabaena variabilis [2Fe-2S] ferredoxin.
The preparation and characterisation of a Ru-modified derivative of the [2Fe-2S] ferredoxin FdI component of A. variabilis by attachment of Ru(NH3)5 to a surface histidine has been carried out. Metal analyses by ICP gave an Fe/Ru ratio of 1.97:1. From NMR the attachment is confirmed as a modification at His-16 and not His-92. No Ru-modification of the [2Fe-2S] FdI component of spinach, which has His-92 but no His-16, was observed. Histidine pKa values have been determined and anomalously low values for A. variabilis (5.3) and spinach (< 5.0) His-92 residues are noted, whereas His-16 gives a pKa of 6.95. Pulse radiolysis techniques with e-aq as reductant have been used to generate the metastable Fe(II)Fe(III)Ru(III) state from Fe(III)2Ru(III), k = 8.8 x 10(10) M-1 s-1. Intermolecular, k(inter) = 4.3 x 10(6) M-1 s-1 at approx. 19 degrees C, but no intramolecular electron-transfer (ET) Fe(II)Fe(III) to Ru(III) contribution was observed with Fe(III)2Ru(III) in the range 7-30 microM. The driving force for ET is approx. 500 mV, and the edge to edge distance from the C gamma of His-16 to the nearest cysteinyl S-atom attached to the Fe2S2 cluster is 16.1 A. Using the Beratan-Onuchic pathway approach, the most favourable ET intramolecular route consists of 20 covalent and 3 H-bonds, with a total distance from the S of Cys-49 to the C gamma of His-16 of 37.1 A. We note that the reduced Fe is on the remote side of the [2Fe-2S] cluster from the site of Ru-attachment.
Lloyd
E
E
Department of Chemistry, University of Newcastle, Newcastle-upon-Tyne, UK.
Tomkinson
N P
NP
Salmon
G A
GA
Sykes
A G
AG
D000595:Amino Acid Sequence; D017033:Anabaena / Q000737:chemistry*; D004579:Electron Transport; D005288:Ferredoxins / Q000737:chemistry* / Q000302:isolation & purification; D006639:Histidine / Q000737:chemistry*; D007501:Iron / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010945:Plants, Edible / Q000737:chemistry; D011675:Pulse Radiolysis; D012413:Rubidium / Q000737:chemistry*
D005288:Ferredoxins; D006639:Histidine; D007501:Iron; D012413:Rubidium