Probing magnetic properties of the reduced [2Fe-2S] cluster of the ferredoxin from Arthrospira platensis by H-1 ENDOR spectroscopy

The H-1 electron nuclear double resonance (ENDOR) spectra in frozen solutio ns of the reduced [2Fe-2S] cluster in ferredoxin from Arthrospira (Spirulin a) platensis have been measured at low temperatures (5-20 K) and simulated using orientational selection methods. The analysis confirmed the existence of a single paramagnetic species with iron valence states II and III conne cted uniquely to the cluster irons. The experimental ENDOR spectra were fit ted to a model including the spin distribution on the centre, the orientati on of the g-matrix, and the isotropic and anisotropic hyperfine couplings o f the nearest protons in the crystallographically determined structure. In order to partially simulate ENDOR line shapes, a statistical distribution o f the corresponding torsion angles between the Fe(III) centre and one of th e beta-CH2 protons was introduced. From the analysis, four of the larger hy perfine couplings found were assigned to the cysteine beta-protons near the Fe(III) ion of the cluster, with isotropic hyperfine couplings ranging fro m 1.6 to 4.1 MHz. The spin distribution on the two iron ions was estimated to be +1.85 for the Fe(III) ion and -0.9 for the Fe(II) ion. The Fe(III) io n was identified as being coordinated to the cysteine ligands Cys49 and Cys 79, confirming previous NMR results. The direction of the g-tensor with res pect to the cluster was deduced. The g(1)-g(2) plane is parallel to the pla nes through each iron and its adjacent cysteine sulfurs; the g(2)-g(3) plan e is nearly perpendicular to the latter planes and deviates by 25 degrees f rom the FeSSFe plane. The g(1) direction is dominated by the bonding geomet ry of Fe(II) and does not align with the Fe(II)Fe(III) vector.