The Mechanism of Folding of Human Frataxin in Comparison to the Yeast Homologue – Broad Energy Barriers and the General Properties of the Transition State

: The funneled energy landscape theory suggests that the folding pathway of homologous proteins should converge at the late stages of folding. In this respect, proteins displaying a broad energy landscape for folding are particularly instructive, allowing inferring both the early, intermediate and late stages of folding. In this paper we explore the folding mechanisms of human frataxin, an essential mitochondrial protein linked to the neurodegenerative disorder Friedreich's ataxia. Building upon previous studies on the yeast homologue, the folding pathway of human frataxin is thoroughly examined, revealing a mechanism implying the presence of a broad energy barrier, reminiscent of the yeast counterpart. Through an extensive site-directed mutagenesis, we employed a Φ -value analysis to map native-like contacts in the folding transition state. The presence of a broad energy barrier facilitated the exploration of such contacts in both early and late folding events. We compared results from yeast and human frataxin providing insights into the impact of native topology on the folding mechanism and elucidating the properties of the underlying free energy landscape. The findings are discussed in the context of the funneled energy landscape theory of protein folding.