Antioxidant activity of sugar-lysine Maillard reaction products in cell free and cell culture systems

The presence of a linear 3Fe-4S] cluster in a protein was first observed in beef-heart aconitase. Here, we report the formation of linear 3Fe-4S] clusters upon guanidine hydrochloride (GuHCl)-induced unfolding of Aquifex aeolicus 2Fe-2S] ferredoxins (Fd) (AaeFd1, AaeFd4, and AaeFd5) at alkaline conditions (pH 10, 20 degrees C). We find the mechanism of linear 3Fe-4S] cluster formation to depend critically on the speed of polypeptide unfolding. In similarity to seven-iron Fds, polypeptide unfolding determines the rate by which linear 3Fe-4S] clusters form in AaeFd4 and AaeFd5. In contrast, in a disulfide-lacking variant of AaeFd1, which unfolds faster than AaeFd4 and AaeFd5, the polypeptides unfold first and the majority of clusters decompose. Next, unfolded polypeptides retaining intact clusters scavenge iron and sulfur to form linear 3Fe-4S] clusters in a bimolecular reaction. Wild-type AaeFd1 unfolds slower than the speed of linear-cluster decomposition, and the linear species is never populated. Linear 3Fe-4S] clusters may be intermediates during folding of iron-sulfur proteins.