Studies of light-induced nickel EPR signals in Desulfovibrio gigas hydrogenase

Abstract

Illumination of the Ni-C state of D. gigas hydrogenase at temperatures below 60 K caused the disappearance of the single Ni-C signal (g= 2.19, 2.14, 2.01) and the simultaneous appearance of two overlapping signals with different g_z and g_y values. The overlapping spectra can be deconvoluted into two independent spectra with g= 2.26, 2.11, 2.044 for the signal here referred to as Ni-C^*₁ and g= 2.29, 2.12, 2.044 for the signal referred to as Ni-C^*₂. The rate of disappearance of the Ni-C signal upon illumination of D. gigas hydrogenase at 5 K was the same as at 30 K, but a complicated new spectrum appeared. This is interpreted as being due to the superposition of the spectra of the reduced [4Fe—4S] clusters and of the two overlapping photoilluminated Ni-C signals which, at this temperature, show splittings due to the interaction with a nearby paramagnet. The splittings are anisotropic. Upon light irradiation of D. gigas hydrogenase at temperature above 60 K, only signal Ni-C^*₂ appeared. This different behaviour of the two overlapping light-induced signals suggests that after illumination of the Ni-C species, two different forms of the Ni site are formed. The differences in the g values indicate slightly different coordination environments. The effects of illumination and temperature suggest that one of them is an intermediate in the formation of the other one, at least at low temperatures. Although irreversible below 100 K, the photolytic process was completely reversible at temperatures above 120 K.