Cytosolic iron-sulfur protein assembly system identifies clients by a C-terminal tripeptide

Abstract

The eukaryotic cytosolic Fe-S protein assembly (CIA) machinery inserts iron-sulfur (Fe-S) clusters into cytosolic and nuclear proteins. In the final maturation step, the Fe-S cluster is transferred to the apo-proteins by the CIA-targeting complex (CTC). However, the molecular recognition determinants of client proteins are unknown. We show that a conserved [LIM]-[DES]-[WF]-COO−tripeptide present at the C-terminus of clients is necessary and sufficient for binding to the CTCin vitroand directing Fe-S cluster deliveryin vivo. Remarkably, fusion of this TCR (target complex recognition) signal enables engineering of cluster maturation on a non-native protein via recruitment of the CIA machinery. Our study significantly advances our understanding of Fe-S protein maturation and paves the way for bioengineering applications.

Trp-ing out on cytosolic [Fe–S]-cluster delivery

“…Because Fe and S are toxic in their free forms, and because [Fe–S]-clusters are labile in aqueous solution, endogenously formed clusters must be protected within protein matrices and trafficked to their cognate clients through specific protein–protein interactions. In PNAS, Marquez et al ( 4 ) describe an important aspect of how [Fe–S]-clusters are delivered to certain client [Fe–S]-proteins produced in the Eukaryotic cytosol.…”

“…A key observation ultimately leading to the discovery of a tripeptide motif in the CIA system, which is analogous to the ISC-associated LYR targeting signal, is that a Trp residue is located at the C-termini of several known CTC targets ( 9 – 11 ). Given that Trp is rarely found at the C terminus of proteins produced in nature, Marquez et al ( 4 ) advanced these observations by performing an exhaustive comparative bioinformatic analysis of the C-terminal region of known or suspected [Fe–S]-proteins, including those found in the cytosol, nucleus, mitochondria, and in the Bacteria and Archaea. Remarkably, a signature tripeptide motif, designated the targeting complex recognition (TCR) signal, having Ile, Leu, or Met as the antepenultimate residue, Asp, Glu, or Ser as the penultimate residue, and Trp or Phe as the C-terminal residue was identified in a subpopulation of [Fe–S]-proteins produced in the cytoplasm or, in some cases, “adaptor” proteins associated with an [Fe–S]-protein produced in the cytoplasm.…”

“…Other intriguing aspects of [Fe–S]-protein maturation have also emerged. A similar approach described by Marquez et al ( 4 ) combining bioinformatic, biochemical, and physiological approaches can now be applied to identify other determinants of client [Fe–S]-protein interactions with the CIA targeting complex and, perhaps, identification of heretofore unknown [Fe–S]-proteins. Also, because defects in the formation or activity of many [Fe–S]-proteins often have pathological consequences ( 2 ), it will be of great interest to determine whether substitutions within the targeting sequence of specific client [Fe–S]-proteins are similarly manifested by a disease state.…”