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2P32 – Principles in Inorganic Chemistry Dr. M. Pilkington

1. What is biological inorganic chemistry (biochemistry)

Lecture 15 – Biological Inorganic Chemistry

2. Functional roles of biological inorganic elements

3. Metal ions and proteins: binding, stability and folding

4. Vitamin B12 - Cobalt an essential element for life

5. Biomineralization

6. Metals in medicine

7. Poisoning by metals

1. What is biological inorganic chemistry (bioinorganic chemistry)?

An interdisciplinary research field at the interface of the more classical areas

of inorganic chemistry and biology/biochemistry.

Understanding the roles that metallic and nonmetallic elements

l i bi l i l s st ms is th l f bi l i l i i play in biological systems is the goal of biological inorganic

(bioinorganic) chemistry.

There are two main fields of bioinorganic chemistry:

1. Investigations of inorganic elements in processes e.g. nutrition, the toxicity of

inorganic species, including the ways in which such toxicities are overcome both

by natural systems and by human intervention, and of metal-ion transport and

storage in biology.

2. The introduction of metals (metal complexes) into biological systems as probes

and drugs.

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The familiar elements C, H, N, O, P and S, the big six, which are well covered in

biochemistry texts provide the major building blocks for cellular components

including proteins, nucleic acids, lipids-membranes, polysaccharides and

metabolites.

Despite this organic diversity life cannot survive with only these principle Despite this organic diversity, life cannot survive with only these principle

elements.

Inorganic elements are also essential to life processes - eleven elements of the

periodic table are required for all forms of life and an additional seven or eight

elements are used by organisms on our planet.

Blood known to contain iron since the 17th century.

Need for Zinc, 1896.

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Transition Elements Relevent to Bioinorganic Chemistry

The Biometals

Why does biology utilize transition metals ?

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2. Functional roles of selected biological inorganic elements Charge balance and electrolytic conductivity: Na, K, Cl

Structure and templating: Ca, Zn, Si, S

Signaling: Ca, B, NO

Bronstead Acid-Base Buffering: P, Si, C

Lewis Acid Base Catalysis: Zn Fe Ni Mn Lewis Acid-Base Catalysis: Zn, Fe, Ni, Mn

Electron Transfer: Fe, Cu,

Group Transfer (e.g. CH3, O, S): V, Fe, Co, Ni, Cu, Mo, W

Redox Catalysis: V, Mn, Fe, Co, Ni, Cu, W, S, Se

Energy Storage: H, P, S, Na, K, Fe

Biomineralization: Ca, Mg, Fe, Si, Sr, Cu, P

Owing to the great advances in research in biological inorganic chemistry we now know the f f h h b l h d d h h l structures of many components of the systems that biology has adapted through evolution

to perform these essential functions. Many relationships between structure and function have been elucidated.

Biological inorganic chemistry has also profoundly impacted both environmental science and medicine

Selected metal ions and their function together with typical

deficiency symptoms

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3. Metal ions and proteins: binding, stability and folding Life has evolved with the minerals of the Earths crust and the ions in the

Earths waters.

Therefore it is not surprising that living beings have evolved the capability touse inorganic elements for key biological processes and to defend themselvesfrom poisoning by other elementsfrom poisoning by other elements.

Some metal ions, when associated with polypeptides, can help catalyze uniquechemical reactions and perform specific physiological functions. We call suchmetal ions “metal cofactors”.

Amino acids and proteins alone are not sufficient to perform all thereactions needed for life. For example, the Fe3+/Fe2+ and Cu2+/Cu+ redoxcouples play critical roles as cofactors for electron transfer reactions in thecatalysis of redox reactionscatalysis of redox reactions.

The Fe2+ ion can reversibly bind dioxygen (O2) if a coordination site is available.

In the periodic table those metal ions essential for life are highlighted ingreen. Some of these e.g. Fe, Cu and Zn are strongly associated with proteinsand form the so-called metalloproteins.

For example, ferritin the metalloprotein that stores iron in the body. In mammals iron is bound and transported by the serum protein transferrin, and

it is stored by ferritin in most life forms. Ferritin is a spherical molecule with an outer coat of protein and an inner core of

hydrous ferric oxide [FeO3(H2O)n]. As many as 4500 atoms of Fe can be stored in a single ferritin molecule.

A three-dimensional representation

showing ferritin, the iron-storage

protein in the body.

Ferritin has a spherical shape, and iron

(brown) is stored as a mineral inside the

sphere.sphere.

We do not yet fully understand the

control of Fe loading during abundance

and mobilization during scarcity.

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Some metal ions are found deeply buried within proteins.

Such metal ions are often “structural” in function.

Their interaction with the protein helps insure the optimal protein structure andcontributes to the stability and appropriate acid-base behavior necessary forthe physiological function.

For example the Zn2+ ions in Zn fingers which are transcription factors are For example, the Zn ions in Zn fingers which are transcription factors arenecessary for the adoption of the proper shape of the protein, which allows it tointeract with DNA. It is not currently known if the zinc ion plays more than astructural role in this proteins i.e. if the Zn2+ concentrations are also used insome manner to regulate gene expression.

Structure of the first zinc finger. Residues 13, 15,

16 and 19 are implicated in DNA recognition, in

this case the base triplet GCG. The zinc ion is in

the lower right portion of the structure and is

chelated by two cysteines and two histidines.

In multicellular organisms, sodium and calcium are found mostly outside thecellular compartment (extracellular), while potassium and magnesium are largelyintracellular.

Calcium and magnesium are often metal activators in proteins to which they bindwith relatively low affinity.

Under appropriate circumstances, these metal ions induce conformationalchanges in the protein upon binding and in doing so they may transmit a signal e.g.the firing of neurons by rapid influx of sodium ions across a cell membrane

Or the regulation of intracellular functions by calcium binding proteins such ascalmodulin.

Structure of calmodulin where all four sites

are occupied with calcium ions, and the linker

has formed a long alpha helix, separating the

two calcium-binding domains.

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4. Vitamin B12 – cobalt an essential element for life.

Cobalt appears centrally in the periodic table, and with its neighbors, iron,

manganese, nickel and copper, has a vital role in a number of biochemical

metalloenzyme reactions.

Vitamin B12, also called cobalamin, is a water-soluble vitamin with a key role in 12, , y

the normal functioning of the brain and nervous system, and for the formation

of blood.

It contains the biochemically rare element cobalt.

It is one of the eight B vitamins.

It is normally involved in the metabolism of every cell

of the human body, especially affecting DNA synthesis

and regulation, but also fatty acid synthesis and

energy production.

Vitamin B12 has a porphryin core:

Porphyrins are heterocyclic macrocycles composed of four modified pyrrole

subunits interconnected at their α carbon atoms via methine bridges.

Porphyrins are aromatic. That is, they obey Hückel's rule for aromaticity,

The macrocycle has 26 π electrons in total.

possessing 4n+2 π electrons (n=4 for the shortest cyclic path) delocalized over

the macrocycle.

Thus porphyrin macrocycles are highly conjugated systems. As a consequence,

they typically have very intense absorption bands in the visible region.

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One of the best-known porphyrins is heme, the pigment in red blood

cells; heme is a cofactor of the protein hemoglobin.

Vitamin B12 was discovered from its relationship to the disease pernicious

anemia, which is an autoimmune disease in which parietal cells of the stomach

responsible for secreting intrinsic factor are destroyed. Intrinsic factor is

crucial for the normal absorption of B12, so a lack of intrinsic factor, as seen in

pernicious anemia, causes a vitamin B12 deficiency.

Medical uses of Vitamin B12

Vitamin B12 is used to treat vitamin B12 deficiency, cyanide

poisoning, and hereditary deficiency of transcobalamin II.

It is given as part of the Schilling test for detecting

In pernicious anemia, the body does not make enough

red blood cells.

pernicious anemia.

High vitamin B12 level in elderly individuals may protect against

brain atrophy or shrinkage associated with Alzheimer's disease

and impaired cognitive function

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Small red crystals of Vitamin B12 were then grown by Lester Smith and given to

Dorothy Hodgkin for X-ray crystal structure analysis. All that was known at this

stage was that the approximate empirical formula was:

C61-64H84-90N14O13-14PCo.

A t l t t l l f thi i d l it h d b A crystal structure on a molecule of this size and complexity had never been

attempted before, it was a huge and complex task, since crystal structure

determinations were not the routine tasks that they are today, and the

techniques were still being developed, both the X-ray and the computer

equipment were tedious and difficult to use. Thus the X-ray crystal structure

which emerged from this study between 1950 the early 1960’s was the first

determination of a chemical formula by X-ray diffraction and the firstdetermination of a chemical formula by X ray diffraction, and the first

determination of the structure of a metalloenzyme.

This achievement is recognized as the birth of Biochemistry.

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It was a triumph for Dorothy

Hodgkin and her Oxford X-ray

crystallography group, inspiring

many young crystallographers, and

pointing them to biochemistry as

an exciting new subject for their

endeavors.

The structure work also caused Woodward (at Harvard) and Eschenmoser (at

the Swiss Federal Institute of Technology) to start synthetic work on Vitamin

B12.

The synthesis took 11 more years and involved more than 90 separate reactions The synthesis took 11 more years, and involved more than 90 separate reactions

performed by over 100 co-workers.

The stereochemical puzzles involved in the synthesis led to the Woodward-

Hoffman rules.

This all adds up to three Nobel prizes in chemistry and one in medicine!

Vitamin B12 is a metalloezyme, about 40% of metalloproteins are

metalloenzymes.

Metalloenzyme- metal ions main role is to function in enzymatic reactions.

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Examples of enzymes classified by metal centres

5. Biomineralization The biological process that give rise to bones, shells and teeth is called biomineralization.

Over the last two decades, the study of biomineralization has shifted more toward a more

chemical perspective and in doing so has become established as a new branch of bioinorganic

chemistry that represents the length scale and interplay between biological processes and

inorganic chemistryinorganic chemistry.

The research aims of biomineralization include the structural and compositional

characterization of biominerals, understanding the functional properties of

biominerals, and elucidation of the processes through which organic macromolecules and

organic structures control the synthesis, construction and organization of inorganic

mineral-based materials. Two examples of biominerals are:

1 C l i Bi i l h ll d i li d ti h b d t th d1. Calcium Biominerals – shells and mineralized tissues, such as bone and teeth are composed

of calcium carbonate or calcium phosphate minerals, combined with a complex organic

macromolecular matrix of proteins, polysacharides and lipids. Calcium carbonate

biominerals such as calcite and aragonite are used for structural support.

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The mother of pearl layer of seashells isa laminate of 0.5-m thick calciumcarbonate (aragonite) polygonal tabletssandwiched between thin 30–nm sheets ifa protein-polysacharide organic matrix.

Calcium Biominerals

The matrix plays a key spatial role inlimiting the thickness of the crystals andis structurally important in themechanical “design” of the shell.

Biominerals also have some unusualfunctions. For example crystals of calciteare used as gravity sensors in a widerange of animalsrange of animals.

The optical properties of calcite areexploited in the lenses of the compoundeyes of extinct creatures called trilobiteswhich are preserved as fossils.

Trilobite – extinct arthropods that dissapeared about 250 million years ago.

The name trilobite mans three lobed since they are made up of three body sections, a longitudinal lobe, a central axis lobe and two symmetrical pleural lobes that flank the axis

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Bone and teeth are made from calcium phosphate in the form of a mineral hydroxyapatite (HAP).

The structure and mechanical properties of bone are derived from the organized mineralization of HAP within a fibrous matrix of a structural protein, collagen along with proteins with sugar sidechains.

The distinction between an inorganic and a bioinorganic mineral is clearly seen The distinction between an inorganic and a bioinorganic mineral is clearly seen in bone, which is close to being described as a “living mineral” since it undergoes continual growth, dissolution and remodeling.

2. Iron Oxides – bioinorganic iron oxides are widespread and serve several functions.

A mixed valence compound magnetite (Fe3O4) is of special biological relevance.

Magnetite is synthesized in a wide range of magnetotactic bacteria.

These organisms are aligned in the Earth’s magnetic field such that in the northern hemisphere they swim downward (north seeking) toward the oxygen depleted zone at the sediment-water interface of fresh water and marine environments.

Magnetic Microbes- magnetotactic bacteria.

Magnetotactic bacteria were discovered in 1975 by Richard P. Blakemore.

Blakemore noticed that some of the bacteria that he observed under a microscope

always moved to the same side of the slide.

If he held a magnet near the slide, the bacteria would move towards the north end

of the magnet. These bacteria are able to do this because they make tiny, iron-

containing, magnetic particles.

Each of these particles is a magnet with a north pole and a south pole. The bacteria

arrange these tiny magnets in a line to make one long magnet. They use this magnet

as a compass to align themselves to the earth's geomagnetic field.

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Why would these bacteria need a compass? Like many other types of bacteria,

magnetotactic bacteria don't like oxygen very much. They will move away from

areas with high oxygen and toward areas with low or no oxygen. In an aquatic

environment, the level of oxygen decreases as one moves deeper into the water.

So, magnetotactic bacteria like to live in the deeper parts of their aquatic g p p q

environments. They use their magnetic compass to tell them which way is down.

Scientists are also interested in practical applications involving these magnetic

microbes.

While it isn't likely we'll be using these bacteria to stick notes to our

refrigerators, they could prove to be useful to humans.

The tiny magnets that these simple organisms make are far superior to those The tiny magnets that these simple organisms make are far superior to those

produced by people. So, scientists and engineers are trying to develop ways to

use this magnetic material in places where tiny magnets are much better than

big magnets.

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6. Metals in Medicine The use of iron and copper can be traced to the ancient Greeks and Hebrews

through their writings. Among metal ions commonly used over the centuries were Hg2+ for the

treatment of syphilis, Mg2+ for intestinal disorders, and Fe2+ for anemia. It is seldom useful to describe elements as “toxic” or “nontoxic”. Even so-

ll d t i d ll b t l t d i l d d hibit called toxic compounds can usually be tolerated in low doses, and may exhibit therapeutic effects within narrow concentration ranges, and biochemically essential elements can be toxic at high doses.

The Betrand diagram schematically summarizes this situation:

All things can be poisons

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Essential element dosage and physiological response

Metal homeostasis - trafficking, and sensing pathways that allow organisms to

maintain an appropriate (often narrow) intracellular concentration range of essential

transition metals.

Dose response – Non essential elements

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Recommended Daily Allowances for Inorganic Elements in the Human Body

Some of the areas of medicinal inorganic chemistry are shown below:

. Today inorganic chemistry is beginning to have a major impact in modern

medicine. Both essential and nonessential metals can be used in therapy and diagnosis.

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Compounds in current clinical use

are summarized to the right.

It is important to ask which parts

of the compound are essential for

activity the metal itself the activity, the metal itself, the

ligands or the intact complex of

metal plus at least some of the

ligands.

Many metallodrugs are “prodrugs”

they undergo ligand substitution

d/ d i b f and/or redox reactions before

they reach the target site.

Metallotherapeutics – cancer is one of the top three killers worldwide and is a

difficult disease to treat. It is hard to find drugs that are both effective and have

low toxicity to the human body as a whole.

Three important inorganic pharmaceuticals are:

1. Platinum drugs such as cisplatin – an anticancer drugm g p g

There is a need to develop new Pt anticancer drugs because cisplatin is a very toxic

compound with severe side effects such as kidney poisoning. Activity is requires

against a wider range of cancer types such as lung, breast and colon cancers. Cancer

cells can also become resistant to cisplatin after repeated treatments. Other drugs

related to cisplatin that are approved for clinical use are carboplatin, and nedaplatin.

Oxaliplatin (trade name Eloxatin) was approved for clinical use in August 2002 forOxaliplatin (trade name Eloxatin) was approved for clinical use in August 2002 for

use in the treatment of metastatic carcinoma of the colon or rectum as a combination

infusion with 5-fluorouracil and leuovorin.

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2. Gold antiarthritic drugs e.g. Auranofin Injectible Au(I) thiolate drugs and one oral Au(I) phosphine drug (Auranofin) are widely

used in clinics today for the treatment of difficult cases of rheumatoid arthritis. There is also interest in the potential use of Au compounds for treating asthma, malaria, cancer and HIV.

Auranofin is thought to deposit Au(I) in the lysosomes (intracellular compartments that house destructive enzymes) and inhibit the enzymes that destroy joint tissues.

There is still much to learn - the cause of rheumatoid arthritis remains unknown to-date.

3. Radiodiagnostic and radiopharmaceutical drugs e.g.Cardiolyte – a heart imaging agent Radionucleotides are used for both imaging and therapy, 99mTc is used in > 85% of all

diagnostic scans in hospitals because of its ideal properties. i.e. it has a half life long enough to allow accumulation in the target tissue, yet short enough to minimize the radiation dose to the patient.

99mTc is a -emitting radionucleotide. Cardiolyte is a +vly charged 99mTc complex Hundreds of isonitrile complexes were Cardiolyte is a +vly charged Tc complex. Hundreds of isonitrile complexes were

investigated to obtain the optimum balance between uptake and clearance in the heart compared to other organs.

The six methoxy ligands are sequentially metabolized in the liver to hydroxyl groups. This transformation turns the complex into increasingly hydrophilic species which are not

retained in myocardial tissue.

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Examples of Metal-based drugs

7. Poisoning by Metals Hg2+, Pb2+ and Ti+ are poisonous by any dose.

Fe and Cu are poisonous in excess (they are excreted, excess only becomes a problem in cases of genetic diseases that affect the excretion of excess.

Fe(toxicity)- inherited – thalassemia (treated with desferrioxamine)

Cu(toxicity) –inherited – Wilson’s disease (British Anti-lewisite helps with this disease it can get rid of copper build up).

See the important ligands in the bioinorganic chemistry handout.

C CCl AsCl

Cl

H H+ BAL C C

Cl

H

H

AsS CH2OH

Lewisite (nerve gas)shuts down respiritorysystem

S

neutralizes the effect of Lewisiteand it can be excreted.