Absorption,transport,storage

1. Absorption,transport,storage Biomineralisation

2. Absorption, transport, storage of metal ions. Biomineralisation (2) Carrying charged ions accross hydrophobic membranes. (3) Transport of ions within the cell and their storage for later use. A balanceddistribution of theelementsinside and outsidethecellsrequires: (1) Mechanismsforselectivecapturingtracequantities of essentialmineralionsintheextracellularenvironment; e.g. solubilisation of mineralprecipitates.

3. Transport and storage of ironin human organism

4. Metabolism of iron 1. Absorption: - Daily iron transport: 10 – 20 mg iron/adult human - Daily absorbed amount:  1 mg Fe/adult human (hemoglobin decomposition: a hem and a globin decomposed and excreted, while most part of the iron is stored in the storing proteins (t1/2  20-30 y) - feedback mechanism: the amount of the absorbed iron is determined by the saturation level of the iron storing proteins. - Site of absorption: duodenum, upper part of small intestine - Affecting factors: pH,solubility, (Fe(OH)3 has very low solubility) hem  Fe(II)  Fe(III) promote:ascorbic acid, citric acid, amino acids (Cys) hinder: stable complexants of iron(III) (polyphenols, e.g. tannins, tee, red wine polyphosphates, e.g. phytic acid in plant seeds) metal ions, Zn(II), Ca(II) (competition at high concentrations)

5. Storage of iron I. The transport/storage of iron in higher organisms are performed by transferrin/ferritine, while in microorganisms by siderophores. Apoferritin: M ~ 450 000, 24 protein subunits (~ 175 amino acids/subunit) diameter: 1300 nm, 2 channels for uptake and release of iron; formed from the hydrophylic and hydrophobic side chains of the protein. Binding of iron: ~ 4500 iron atoms/ ferritin  (~1 Fe/1 amino acid !, 25% iron content) ~ 700 nm d iron containing micelle approx. composition: (FeOOH)8.FeO.H2PO4 oxo-, hidroxo-bridged iron(III)octahedrons strong antiferromagnetic coupling between the iron(III) ions phosphate: „cover layer"  link between the iron core and the protein uptake of iron: in the form of iron(II), then oxidation to iron(III) Hemosiderin: it functions in case of iron overload iron(III)-oxide-hidroxide-phosphate - less ordered - higher iron(~ 40 %) andphosphate content

6. Storage of iron II. Schematic structure of apoferritin Structure of ferritin from protein crystals

7. Storage of iron III. Channels for uptake/release of iron in ferritin

8. Electronmicroscopic picture of ferritin The more saturated the protein protein-hollow the more regular octahedral structures are the iron(III)-oxide-phosphate clusters.

9. Transferrins I. • Transferrins (ovotransferrin, lactoferrin and serum transferrin) are  8 kDa molecule mass glycoproteins, • They consist of two subunits, 1-1 Fe binding sites (log K ~ 22) • Binding site: 2 Tyr-O-, 1 His-N, 1 Asp-COO-, 1 bidentate carbonate (in H-bonding with Arg and Thr side chains and 2 peptide-NH groups) • Fe2+ + HCO3– + Tf = Fe3+-Tf-CO32– + e– + 3 H+ • The Fe3+ reaches the cell through the membrane by receptor mediated endocytosis: cellmembrane pH > 5.5 pH < 5.5

10. Transferrins II. (structure of human lactoferrin) It consists of two subunits, each of them containing 1 iron atom.

11. Transferrins III. (ironbinding site in human lactoferrin)

12. Absorption, transport and storage of copper Metabilitic processes of copper is much less explored than that of iron. This might be explained by the less amount of the metal in the organisms and its many different functions. 1. Absorption and transport of copper: Absorption of copper occurs in the form of CuII in the GI tract in lmm amino acid complexes and reaches the circulatory system bound to human serum albumin. It is transported to the liver by albumin, where ceruloplasmin is synthesised and bound to this protein copper partly get back to the circulatory system. stomach(CuII) → circulation [CuII(His)2 → CuII-albumin] → liver [CuII-ceruloplasmin → CuI-metallothionein] → circulation [CuII-ceruloplasmin + CuII-albumin] → chaperonok → cells [CuI/II-containing enzymes]

13. Distribution of copperinthecirculation: ~ 0,1 % inCu(His)2complex ~ 5-10 % inCu(II)-albumincomplex ~ 90-95 % boundtoceruloplasmin. Basedonthesedataceruloplasminwasconsideredearlierasthecoppertransporter, but more recentdatapointtotherole of albumin. Albumin bindscopperunusuallyin an oligopeptide-likemanneratthe N-terminus. Thisbindingmode has hightermodynamicstability butkineticallylabile, incontrastwiththe inert copperceruloplasminbond. Human albumin: AspAlaHis......... (HIS atposition 3 providesextremestability.) Dog albumin: GluAlaTyr..... (The „coppertolerability” of dogs is significantlylowerthanthat of humen)

14. Albumin is the primary copper transporter for the cells. However, other proteins may also play important roles in transferring copper accross cell membranes and transporting copper in the cells. They are called as „copper-chaperons”, which are specific and usually Cys rich copper transporter proteins. (Similar roles are assumed in case of the prion proteins.) 2. Storage of copper Copper is stored mostly in the liver (spleen, bile). The „cuprein” proteins had been considered earlier as copper stores, but more recent results point to the role of certain enzymes, e.g. erythro- cuprein = CuZn-SOD. Today it is thought that metallothioneins are the primary copper storage proteins. Thionein: low molecular mass Cys rich proteins (polypeptides) extreme high soft metal ion affinity.

15. Metallothioneins A metallothioneins occur in humen, in animals and plants (phytochelatins), they are low molecular mass (6-7 kDa) proteins, which bind soft metal ions (CuI, ZnII, CdII, Hg2II, HgII, AgI és CoII) in cluster structure. Their sulphur and metal contents are very high, may reach 10%. Generally they consisit of two clusters (3M-3S és 4M-5S), in which the metal ions coordinate through Cys-thiolates. The polypeptid part features repeated Cys-X-Cys sequents, in which X stands for a non-Cys amino acid. In the middle of the Figure 12 terminal and 8 bridging CYS side chain bind all together 7 Cd2+-ions, in a chair conformation [3M-3S] cluster (Cd3S9) and an adamantane conformation [4M-5S] cluster (Cd4S11).

16. Metallothioneins • Their basic functions depend on the organism and the peptide variants: • As metal storing proteins they participate in the homeostasis of metal ions first of all that of copper and zinc. • As detoxification molecules they are active in the removal of detrimental soft metal ions (such as CdII, HgII, AgI and AuI). • Their synthesis is induced by some essential, Zn and Cu, but some toxic metal ions, Cd, too. • Inorganic-Hg does, but organic-Hg does not induce formation of metallothioneins.

17. Calcium binding proteins Extracellular proteins: Osteocalcin plays a role in mineralisation of bones Structure of bones: Ca2+, PO43- the main inorganic components of bones: Ca10(PO4)6(OH)2 Other constituents: Mg2+, Na+, CO32-, Cl-, F-, citrate, other anions

18. Mineralisation of bones Components of bones:Ca2+, PO43- main constituents of bones: Ca10(PO4)6(OH)2 otherions: Mg2+, Na+, CO32-, Cl-, F-, citrate, otheranions Ca2+ accumulatesinthecalcificationcells „vesicle" activation of ATPase, pyrophosphataseconcentration ofPO43-increases [Ca2+]3[PO43-]2 > L (precipitation) Role of collagenasmatrixmaterial

19. Mineralisation of bones

20. x H4SiO4 [SiO4]4- The processes of siliciphication Condensation may occur with alcoholic-OH groups too: Diatoma Formation of esters between silicic acid and serin

21. Processes of siliciphication

22. Processes of siliciphication

24. Ellenőrző kérdések Jellemezze a vas anyagcseréjét! Milyen metallo-proteinek vesznek részt benne? Hasonlítsa össze a transzferrin, a ferritin, a chaperonok és a metallothioneinek fémion kötését szerkezeti, termodinamikai és kinetikai szempontból! Változott-e a létfontosságú elemek csoportja a kémiai és biológiai evolúció során? Példákkal igazolja állítását! Milyen fontosabb biomineralizációs folyamatokat ismer? Jellemezze a csontképződés folyamatát! Mi az a szilicifikációs folyamat és hol van jelentősége?