title: Chemical rescue of a site-modified ligand to a [4Fe-4S] cluster in PsaC, a bacterial-like dicluster ferredoxin bound to Photosystem I.
abstract: Chemical rescue of site-modified amino acids using externally supplied organic molecules represents a powerful method to investigate structure-function relationships in proteins. Here we provide definitive evidence that aryl and alkyl thiolates, reagents typically used for in vitro iron-sulfur cluster reconstitutions, serve as rescue ligands to a site-specifically modified [4Fe-4S](1+,2+) cluster in PsaC, a bacterial dicluster ferredoxin-like subunit of Photosystem I. PsaC binds two low-potential [4Fe-4S](1+,2+) clusters termed F(A) and F(B). In the C13G/C33S variant of PsaC, glycine has replaced cysteine at position 13 creating a protein that is missing one of the ligating amino acids to iron-sulfur cluster F(B). Using a variety of analytical techniques, including non-heme iron and acid-labile sulfur assays, and EPR, resonance Raman, and Mössbauer spectroscopies, we showed that the C13G/C33S variant of PsaC binds two [4Fe-4S](1+,2+) clusters, despite the absence of one of the biological ligands. (19)F NMR spectroscopy indicated that the external thiolate replaces cysteine 13 as a substitute ligand to the F(B) cluster. The finding that site-modified [4Fe-4S](1+,2+) clusters can be chemically rescued with external thiolates opens new opportunities for modulating their properties in proteins. In particular, it provides a mechanism to attach an additional electron transfer cofactor to the protein via a bound, external ligand.
authors: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA. antonkin@physik.fu-berlin.de
authors: Mikhail L
authors: ML
authors: Antonkine
authors: Estelle M
authors: EM
authors: Maes
authors: Roman S
authors: RS
authors: Czernuszewicz
authors: Christoph
authors: C
authors: Breitenstein
authors: Eckhard
authors: E
authors: Bill
authors: Christopher J
authors: CJ
authors: Falzone
authors: Ramakrishnan
authors: R
authors: Balasubramanian
authors: Carolyn
authors: C
authors: Lubner
authors: Donald A
authors: DA
authors: Bryant
authors: John H
authors: JH
authors: Golbeck
mesh_terms: D005288:Ferredoxins / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000378:metabolism*; D008024:Ligands; D045331:Photosystem I Protein Complex / Q000737:chemistry / Q000378:metabolism*
chemical_list: D005288:Ferredoxins; D007506:Iron-Sulfur Proteins; D008024:Ligands; D045331:Photosystem I Protein Complex; C089909:photosystem I, psaB subunit