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Chapter 8: Biopolymers Examples of biopolymers are: Starch Cellulose Proteins Nucleic Acids Polymer Nylon 6 PS PBI Modulus 1.5 GPa 3 GPa 6 GPa Strength 36 MPa 45 MPa 186 MPa Biopolymers’ locations Animal Cell Plant Cell RNA Nucleus Cytoplasm1 Nucleus Cytoplasm DNA Nucleus Organelles (e.g. Mitochondrion) Nucleus Organelles (e.g. Chloroplast) Starch - Chloroplasts Cellulose - Cell Walls Fish Red Blood Cell Note: Not an exhaustive list, these are a few representative examples ~ 15m 1 Cytoplasm: the organic and inorganic material inside the cell but outside its nucleus. Deoxyribonucleic Acid DNA Tensile 476±84 pN isolated during war in 1860’s in puss from wounds 3.14 x 10-18m2 and 4.76 x 10-10 N 1.5 x 108 Pa 2 nm The Human genome (all the nuclear DNA) has approximately 3 x 109 nucleotide monomers in the shape of a double helix with a radius of ~ 1 nm. Chromatin packing of DNA A B 2 nm 2 nm 1111 nmnm D 300300 nm nm E 700 nm 700 nm F C 3030 nmnm 1400 1400 nmnm Upon “melting” DNA strands can be replicated RNA is less stable & is never found in old bones Photocrosslinking leads to a helix that won’t un-zip!! DNA Melting Proteins O H2N R Insulin crystal Strong inter- and intra-molecular effects beta sheets alpha helices N H OH O O H2N Proteins by Function R N H OH O O H2N R N H OH O O H2N R N H OH O O H2N R N H OH O O H2N R N H OH O Proteins O H2N R N H OH O • The control of protein structure builds information into the molecule that translates into function • Proteins are the most common biological macromolecules in the extra cellular matrix • Perform structural and functional tasks – Collagen (triple helix – gly-X-Y) where proline and hydroxy proline is often present is the basic stuctural protein – Enzymes perform specific catalytic tasks – Adhesive proteins are bind cells to substrates – fibronectin, integrin, etc. – Provide signal transduction between cells and ECM Protein Structure O Primary - identitiy and order of amino acids H N N H R -determines all other levels of structure -covalent bonding Secondary - helices % sheets, turns, random coils -driven & stabilized by hydrogen bonding -sterics Tertiary - 3-D Folded structures -hydrophobic interactions -often direct determinant of function Quaternary - multiple peptides aggregating -multiple bonding interactions OH 2 O Structure is a consequence of sequence Function is a consequence of structure Primary Structure: Amino Acid Sequence •20 amino acids • O R2 H2N N H R1 Alanine Arginine O H2N CHC OH CH3 Asparagine O H2N CHC OH H2C H2C H2C HN C NH NH2 Glycine O H2N CHC OH CH2 C O NH2 Histidine N NH Proline H N C O OH O H2N CHC OH H2C C O OH Isoleucine O H2N CHC OH CH2 O H2N CHC OH H Aspartic Acid O H2N CHC OH CHCH3 H2C CH3 Serine O H2N CHC OH CH2 OH Leucine O H2N CHC OH CH2 CHCH3 CH3 Threonine O H2N CHC OH CHOH CH3 O H N O R3 R4 N H O H N O OH R5 Cysteine Glutamic acid O H2N CHC OH CH2 SH O H2N CHC OH H2C H2C C O OH Lysine O H2N CHC OH H2C H2C H2C H2C NH2 Methionine Phenylalanine O H2N CHC OH CH2 CH2 S CH3 O H2N CHC OH H2C Tryptophan O H2N CHC OH CH2 Tyrosine O H2N CHC OH CH2 HN OH Glutamine O H2N CHC OH CH2 CH2 C O NH2 Valine O H2N CHC OH CHCH3 CH3 Primary Structure: Amino Acid Sequence •20 amino acids • O H2N R1 R2 N H O O H N R3 R4 N H O H N O OH R5 Average protein 300-400 amino acids = 30-45K Daltons Protein with 300 mers based on 20 amino acids: P =20300 or 10390 different possible sequences Estimated: 100,000 human proteins (coded by 30,000 genes) Identified: 10,000 human proteins O H2N H N H H O H2N Me O Me N H O H N O N H H H N H O Me O H N O Me N H H HN H O O H N O O N H H N O N H O H N HN Me H H Me O H3N O H N Me N H N H meta-enkephalin O H N O H N Polyglycine H OH Me O O O S HO OH O Me O O H N Me Polyalanine Primary Structure: Amino Acid Sequence •20 amino acids • All one stereoconfiguration Two configurations possible Only (S)-isomers of amino acids used in life on Earth Mirror images Secondary Structure: Helix Pauling 1954 Nobel Prize 3.6 aa per turn Alanine Methionine Glutamate Sheet Valine Leucine Tyrosine Beta- or Hair-Pin Turn two to five residues, of which one is frequently a glycine or a proline Secondary Structure: Random Coiling Secondary Structures: Tertiary Structure: Folding Quaternary Structure: Aggregation Tetramers haemoglobin Quaternary Structure: Coils Eg. Collagen Quaternary Structure: Dimers Eg. Collagen Catabolic activator protein Quaternary Structure: Complex Eg. Collagen Catabolic activator protein Prostaglandin H2 synthase-1 Protein Structure Overview Prions normal abnormal Denaturation loss of 3-D conformation by heat, pH, organic solvents, detergents Spiders spin 6 different fibers Flagelliform Large diameter egg Case fiber (Tubuliform) Aggregate Tubuliform Minor ampullate Capture Spiral (Flagelliform) Pyriform Major ampullate Aciniform Glue coating (Aggregate silk) (?) Wrapping and egg case fiber (aciniform) Web reinforcement (Minor ampullate 1 and 2) Dragline (major ampullate 1 and 2) Vollrath, F. J. Biotechnol. 2000, 74, 67-83. Hu, X. et al. Cell. Mol. Life Sci. 2006, 63, 1986-1999. Pyriform silk (?) The classic strong synthetic fiber Kevlar®: Dupont (1960s) Uses - Bulletproof vests and helmets - Automobile brake pads - Ropes and cables - Aerospace components Material Dragline Silk Kevlar Rubber Nylon, type 6 Fiber axis Strength (GPa) Elasticity (%) Energy to break (J/kg) 1.1 3.6 0.001 0.07 35 5 600 200 5 4 x 10 4 3 x 10 4 8 x 104 6 x 10 Lewis, R. Chem. Rev. 2006, 106, 3762-3774. Vollrath, F.; Knight, D.P. Nature 2001, 410, 541-548. Tanner, D.; Fitzgerald, J.A.; Phillips, B.R. Angew. Chem. Int. Ed. Engl. Adv. Mater. 1989, 5, 649-654. Kubik, S. Angew. Chem. Int. Ed. 2002, 41, 2721-2723. Spider silks have potential in many applications Biomedical applications Surgical sutures Scaffolds for tissue engineering Technical and industrial applications High strength ropes/cables Ballistics Parachutes Fishing line Forced silking to obtain silk fibers Spiders are anesthetized with CO2 and secured ventral side up Silk is pulled from the spinneret, attached to a reel, and drawn at a specified speed Work, R. W.; Emerson, P. D. J. Arachnol. 1982, 10, 1-10. Elices, M.; Perez-Rigueiro, J.; Plaza, G. R.; Guinea, G. V. JOM 2005, 57. Proposed secondary structure and mode of elasticity • Poly(Ala) modules form anti-parallel β-sheets (~30-40%) • Glycine-rich, amorphous regions are thought to be helical Crystalline region with -sheet structure Strain Disordered chain region Kubik, S. Angew. Chem. Int. Ed. 2002, 41, 2721-2723. Van Beek, J. D.; Hess, S.; Vollrath, F. Meier, B. H. Proc. Nat. Acad. Sci. 2002, 99, 10266-10271. Primary structure of spider dragline silk Fibrous protein composed of Spidroin 1 (MaSp1) and Spidroin 2 (MaSp2) - Sequences highly conserved - Repetitive stretches of poly(Ala) and (GlyGlyXaa)n sequences (Xaa = Tyr, Leu, Gln) - MW of MaSp1 ~ 275-320 kDa; Sp1+Sp2 ~ 700-750 kDa Repeating sequence of MaSp1 QGAGAAAAAAGGAGQGGYGGLGGQGAGQGGYGGLGGQGAGQGAGAAAAAAAGGAGQGGYGGLG GLGGYGGQGAGGAAAAAAGAGQGGRGAGQS SQGAGRGGLGGQGAGAAAAAAAGGAGQGGYGGLG GLGGYGGQGAGGAAAAAAGQGGRGAGQN SQGAGRGGLGGQAGAAAAAAGGAGQGGYGGLGGQGAGQGGYGGLG GLGGYGGQGAGGAAAASAGAGQGAGQGGLGGQGAGGAAAAAAAGAGQGGLGGRGAGQS SQGAGRGGEGAGAAAAAAGGAGQGGYGGLGGQGAGQGGYGGLG GLGGYGGQGAGGAAAAAAGAGQGAGQGGLGGQGAGGAAAAGAGQGGLGGRGAGQS SQGAGRGGLGGQGAGAVAAAAGGAGQGGYGGLG GLGGYGRQGAGGAAAAAAGAGQGGRGAGQS NQGAGRGGLGGQGAGAAAAAAAGGAGQGGYGGLG GLGGYGGQGAGGAAAAAGQGGRGAGQN SQGAGRGGQGAGAAAAAAVGAGQEGIRGQGAGQGGYGGLG GAGGYGGQRVGGAAAAAAGAGQGAGQGGLGGQGAGGAAAAAAGAGQGGLGGRGSGQS SQGAGRGGQGAGAAAAAAGGAGQGGYGGLGGQGVGRGGLGGQGAGAAAAGGAGQGGYGGVG SSLRSAAAAASAASAGS Hinman, M.B.; Jones, J. A.; Lewis, R. TIBTECH 2000, 18, 374-379. Vollrath, F.; Knight, D. P. Nature 2001, 410, 541-548. Simmons, A. H.; Michal, C. A.; Jelinski, L. W. Science 1996, 271, 84-87. BioSteel® - Genetically modified goats produce silk in mammary glands - Silk is spun from the goats’ milk Extrusion through “spinnerets” produces fibers Aqueous spinning process is environmentally friendly - Anticipated uses: Surgical sutures Adhesives Fishing line Body armor/military applications Lazaris, A. et al. Science 2002, 295, 472-476. Karatzas, C. N.; Turcotte, C. 2003, PCT Int. Appl. WO03057727. Karatzas, C. 2001, PCT Int. Appl. WO0156626. Islam, S. et al. 2004, U.S. Pat. 20040102614. Proline & Glycine Tensile Strength 1-7 MPa Modulus 10 MPa Carbohydrate Polymers amylopectin •Constituent in starch •Plants store energy •Animals use glycogen (106 glucose) ~10,000 glucose units Polysaccharides • Polymers composed of sugars • Similar to synthetic polymers in that primary structure, DP not as fixed as proteins • Uses include energy storage, component of extra cellular matrix (hyaluronan) Cellulose Animal enzymes ineffective Tensile Strength: 800 MPa •Structural plant material Modulus 75-100 GPa •Cotton = >95% cellulose, wood = 50% •MW of cellulose in 400,000 g/mol, corresponding to about 2200 D-glucose units per molecule •Stiff rods Conformation •Well-organized water-insoluble fibers (20 nm diameter, 40,000 nm long) •The -OH groups form numerous intermolecular hydrogen bonds adding strength to the network. •70% crystalline structure •Tg = 227 °C, TB = 298 °C Cellulose: Fibrous structure Strong hydrogen bonding interactions ( 35 kJ/mole) from 3 hydroxyl groups per sugar monomer Acidic Polysaccharides Acidic polysaccharides are a group of poly saccharides that contain carboxyl groups and/or sulfonic esters. These compounds play an important roles in structure and function of connective tissues. These tissues form the matrix between organs and cells that provides mechanical strength as well filtering the flow of molecular information between cells. Many connective tissues are made up of collagen, a structural protein, in combination with an assortment of acidic polysaccharides that interact with collagen to form loose or tight networks. OSO3- * O HO D-glucuronic acid O O HN O OH O O CH3 O HO N-acetyl-D-glucosamine OH O O -O L-iduronic acid O 3S HN O O SO3O HO HO OSO3 D-glucosamine -OOC OSO3O NH O SO3- D-glucosamine Pentasaccharide unit of heparin responsible for binding to antithrombin III. * Hyaluronic acid D-glucuronic acid N-acetyl-D-glucosamine O OH O O * HO HO O 1 OH 2 3 O 1 O NH O * CH3 Hyaluronic acid is the simplest acidic polysaccharide present in connective tissue MW of ~ 105 and 107 g/mol and contains 30.000 to 100,000 Found in embryonic tissues and specialized connective tissues such as synovial fluid, the lubricant of joints in the body, and the vitreous humor of the eye where it provides a clear, elastic gel that maintains the retina in proper position. Cellulose pulp Enzymes Alkalies Enzymatic processing Alkali cellulose CS2 NaOH ksantogenation Cellulose xanthate Dissolving Ripening Spinning acidic bath Fibre Other products Similar in both processes Waste Na2SO4 / OH2O Spinning acidic bath Fibre Other products acetate rayon O OH O HO O O O OH + O 3 H3C O H3C CH3 O O acetic anhydride glucose unit in cellulose fiber O O O CH3 O O H3C O A fully acetylated glucose unit Tensile 10-250 Mpa Modulus 2 GPa Viscose rayon S C S NaOH Cellulose OH Cellulose O-Na+ S Cellulose O C S-Na+ Na+ salt of a xanthate ester