CHARACTERIZATION OF A FERREDOXIN FROM DESULFOVIBRIO-VULGARIS (HILDENBOROUGH) THAT INTERACTS WITH RNA

The purification and characterization of a ferredoxin from Desulfovibr io vulgaris (Hildenborough) is described. The protein can be isolated in two forms; the major form is strongly complexed to RNA, while a min or form is free from nucleic acid. Bound RNA cannot be removed by dige stion with nucleases, or by heating to 70 degrees C, and it can only b e partially removed by rechromatography. The ultraviolet/visible spect rum shows typical absorption maxima at 280 nm and 400 nm for the RNA-f ree ferredoxin. The RNA-bound protein exhibits an additional strong pe ak at 260 nm. The RNA can be extracted from the protein with phenol Th e ferredoxin is a dimer of subunits, each of 7.5 kDa; its pi is 3.9. T he protein contains a [4Fe-4S]((2+;1+)) cluster with an EPR spectrum ( g = 1.90, 1.93 and 2.05) in the reduced state. A reduction potential o f -360 mV was determined for the RNA-free ferredoxin with reversible v oltammetry at glassy carbon. From the temperature dependence of the re duction potential, the unusually high standard reaction entropy was ca lculated as Delta S-degrees = -230 J . K-1. mol(-1). No electrochemica l response was obtained with the RNA-bound ferredoxin. Binding of RNA appears to require the presence of an intact cluster, since the absenc e of absorption at 400 nm runs in parallel with the absence of absorpt ion at 260 nm. The possibility is discussed that the binding to the RN A has a regulatory function and is controlled by the state of the clus ter.