Cysteine as a ligand platform in the biosynthesis of the FeFe hydrogenase H cluster.
Hydrogenases catalyze the redox interconversion of protons and H2, an important reaction for a number of metabolic processes and for solar fuel production. In FeFe hydrogenases, catalysis occurs at the H cluster, a metallocofactor comprising a [4Fe-4S]H subcluster coupled to a [2Fe]H subcluster bound by CO, CN(-), and azadithiolate ligands. The [2Fe]H subcluster is assembled by the maturases HydE, HydF, and HydG. HydG is a member of the radical S-adenosyl-L-methionine family of enzymes that transforms Fe and L-tyrosine into an [Fe(CO)2(CN)] synthon that is incorporated into the H cluster. Although it is thought that the site of synthon formation in HydG is the "dangler" Fe of a [5Fe] cluster, many mechanistic aspects of this chemistry remain unresolved including the full ligand set of the synthon, how the dangler Fe initially binds to HydG, and how the synthon is released at the end of the reaction. To address these questions, we herein show that L-cysteine (Cys) binds the auxiliary [4Fe-4S] cluster of HydG and further chelates the dangler Fe. We also demonstrate that a [4Fe-4S]aux[CN] species is generated during HydG catalysis, a process that entails the loss of Cys and the [Fe(CO)2(CN)] fragment; on this basis, we suggest that Cys likely completes the coordination sphere of the synthon. Thus, through spectroscopic analysis of HydG before and after the synthon is formed, we conclude that Cys serves as the ligand platform on which the synthon is built and plays a role in both Fe(2+) binding and synthon release.
Department of Chemistry, University of California, Davis, CA 95616;
Daniel L M
DL
Suess
Department of Chemical Engineering, Stanford University, Stanford, CA 94305;
Ingmar
I
Bürstel
Department of Chemical Engineering, Stanford University, Stanford, CA 94305;
Liliana
L
De La Paz
Department of Chemistry, University of California, Davis, CA 95616;
Jon M
JM
Kuchenreuther
Department of Chemistry, University of California, Davis, CA 95616;
Cindy C
CC
Pham
Department of Chemistry, University of California, Davis, CA 95616;
Stephen P
SP
Cramer
Department of Chemical Engineering, Stanford University, Stanford, CA 94305; Department of Bioengineering, Stanford University, Stanford, CA 94305.
James R
JR
Swartz
Department of Chemistry, University of California, Davis, CA 95616; rdbritt@ucdavis.edu.
R David
RD
Britt
D002384:Catalysis; D020134:Catalytic Domain; D003545:Cysteine / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D029968:Escherichia coli Proteins / Q000737:chemistry*; D006864:Hydrogenase / Q000378:metabolism; D007501:Iron / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D008024:Ligands; D008715:Methionine / Q000737:chemistry; D011190:Potassium Cyanide / Q000737:chemistry; D011485:Protein Binding; D011522:Protons; D012993:Solar Energy; D015534:Trans-Activators / Q000737:chemistry*; D014443:Tyrosine / Q000737:chemistry
D029968:Escherichia coli Proteins; D007506:Iron-Sulfur Proteins; D008024:Ligands; D011522:Protons; D015534:Trans-Activators; C423908:zraR protein, E coli; D014443:Tyrosine; D008715:Methionine; D007501:Iron; D006864:Hydrogenase; D003545:Cysteine; D011190:Potassium Cyanide
FeFe hydrogenase; HydG; metallocofactor biosynthesis