Principles of Bioinorganic Chemistry - 2003

1. Principles of Bioinorganic Chemistry - 2003

2. Principles of Bioinorganic Chemistry Two Main Avenues of Study • Understand the roles of naturally occurring inorganic elements in biology. By weight, > 50% of living matter is inorganic. Metal ions at the core of biomolecules control many key life processes. • Use metals as probes and drugs Examples: Cisplatin, auranofin as pharmaceuticals Cardiolyte (99mTc) and Gd, imaging agents

3. Enterobactin: a Bacterial Siderophore

4. Enterobactin, a Cyclic Triserine Lactone A specific cell membrane receptor exists for ferric enterobactin. Release in the cell can occur by hydrolysis of the lactone, reduction to Fe(II), and/or lowering the pH.

5. Structure of Vanadium(IV) Enterobactin

6. Scheme showing the ATP-driven uptake of ferric enterobactin into E. coli cells through a specific receptor in the cell membrane. Does not distinguish D from L outer membrane cytoplasmin membrane intracellular esterase; hydrolyzes Ent, releases iron See Raymond, Dertz, and Kim, PNAS, 100, 3584.

7. Control and Use of Metal Ion Concentrations PRINCIPLES: • Homeostasis: maintain [M+] in proper range • Detoxification: remove excess and/or unnatural metal ions • Extracellular carriers • Passive transport • Ion channels/pumps • Metalloregulation • Binding and release of metal ions to receptors controlled by pH and redox changes • Ion concentration gradients - used to transmit energy and information

8. Note hinge motion that accompanies iron/carbonate binding

9. Transferrin and Structural Changes on Fe Binding Baker, Anderson, and Baker, PNAS, 2003, 100, 3579.

12. Various Anions Can Bind Transferrin Nomenclature: Fbp, ferric binding proteins n, for Neisseria meningitidis Iron must bind as Fe(III), or the ferric state. If reduced, a bacterial reductase must be involved, thus affording control of iron binding and uptake in the organism (see E1/2 values in the table above. Crumbliss, et al. PNAS, 2003, 100, 3659.

13. Mechanism of Transferrin Uptake and Iron Release in Cells by Receptor-Mediated Endocytosis

14. Metal Regulation of Gene Expression PRINCIPLES: • Homeostasis: maintain [M+] in proper range • Detoxification: remove excess and/or unnatural metal ions • Metal-mediated protein structure changes affect transcription • Metal-mediated protein structure changes affect translation • Metal-induced protein structure changes also activate enzymes ILLUSTRATIONS: • Iron regulatory proteins (IRPs); control Ft and Tf translation • Regulation of a toxic metal, mercury • Zinc finger proteins control transcription • Ca2+, a second messenger and sentinel at the synapse

15. Regulation of Iron Levels in Cells The Players: • Ferritin, the iron storage protein: 24-subunits, ~175 aa each; has cubic symmetry; apoFt can house 1000 iron atoms in its central core; a ferroxidase center loads the iron into the protein • Transferrin, the uptake protein, discussed previously Metalloregulation: • In bacteria, occurs at the transcriptional level • In mammals, the synthesis of apoferritin and of the transferrin receptor are regulated at the level of translation, not transcription Central dogma of molecular biology: DNA mRNA Protein transcription translation

16. Ferritin Subunit and Channel Structure

17. Mixed-valent polyiron oxo cluster prepared as a model for ferritin core formation intermediates. Taft, Papaefthymiou, & Lippard, Science 1993, 259, 1302 Overall formula: [Fe12O2 (OCH3)18(O2CCH3) 6(CH3OH)n]

18. Reminder: Apo (left) and Holo (right) Forms of Transferrin Only Iron-Loaded Transferrin Binds to the Receptor

19. Metalloregulation of Iron Uptake and Storage Bacteria: A single protein, Fur (for iron uptake regulator), controls the transcription of genes involved in siderophore biosynthesis. Fur is a dimer with subunits of Mr 17 kDa. At high iron levels, the Fur protein has bound metal and interacts specifically with DNA repressing transcription. Mammals: Expression of ferritin and the transferrin receptor is regulated at the translational level.

20. Components of the Metalloregulatory System IRP Iron-responsive protein (IRP) Stem-loop structure in the mRNA IRP

21. Regulation events High Fe, low TfR, high Ft Low Fe, high TfR, low Ft Fe IRP Message translated Message degraded Ferritin Transferrin IRP Message blocked Message translated

22. IRP1 is the Cytosolic Aconitase Contains an Fe4S4 Cluster Cluster assembled in protein, which then dissociates from mRNA Apoprotein stays associated with mRNA

23. Regulation of a Toxic Metal, Mercury The problem: Mercury in the environment of industrial plants is converted by bacterial to harmful organomercury compounds. Fish and other plant and animal life assimilate the mercury which ultimately enters the human food chain. Bacteria defend themselves against the mercury by using the proteins listed below. The players: Organomercurial lyase Mercuric ion reductase MerR, the intracellular mercuric ion sensor The implications: Transcription of the genes encoding the proteins is controlled by MerR in response to mercury levels

25. Postulated Mechanism for Organomercurial Lyase

26. MerR and Mercuric Ion Reductase Properties Reductase: no structural or detailed mechanistic information MerR EXAFS spectroscopy and chemical modification experiments indicate that Hg-MerR has a 3-coordinate, Hg(S-Cys)3 environment with an average Hg–S distance of 2.43 Å. This unusual tridentate heavy metal receptor site is consistent with the thermodynamic stability of [Hg(SR) 3]- complexes and may account both for the high affinity of the Hg(II) binding and for the selectivity for Hg(II) over other soft metal ions that prefer tetrahedral metal-thiolate coordination.

27. Effect of [Hg2+] on Transcription Activity