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 DNA Nucleus Cytoplasm 1 Nucleus Organelles (e.g. Mitochondrion) Nucleus Cytoplasm Nucleus Organelles (e.g. Chloroplast) Fish Red Blood Cell Starch Chloroplasts Cellulose Cell Walls Note: Not an exhaustive list, these are a few representative examples 1 ~ 15 m 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 -18 m 2 1.5 x 10 8 Pa and 4.76 x 10 -10 N 2 nm

The Human genome (all the nuclear DNA) has approximately 3 x 10 9 nucleotide monomers in the shape of a double helix with a radius of ~ 1 nm.

A B C

Chromatin packing of DNA

D E F

700 nm 700 nm

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

Insulin crystal

Proteins

O H 2 N R N H O OH Strong inter- and intra-molecular effects beta sheets alpha helices

Proteins by Function O H 2 N R N H O OH

O H 2 N R N H O OH

O H 2 N R N H O OH

O H 2 N R N H O OH

O H 2 N R N H O OH

Proteins

O H 2 N R N H O OH • 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

Primary

- identitiy and order of amino acids H 2 N -determines all other levels of structure -covalent bonding

Secondary

- helices % sheets, turns, random coils R O N H -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 O OH

Structure is a consequence of sequence Function is a consequence of structure

Primary Structure: Amino Acid Sequence

Alanine O H 2 N CHC CH 3 OH Glycine O H 2 N CHC H OH •20 amino acids • Arginine O H 2 N CHC H 2 C H 2 C H 2 C HN C NH NH 2 OH Asparagine O H 2 N CHC CH 2 C O NH 2 OH H 2 N R 1 O N H R 2 O H N O R 3 N H R 4 O H N R 5 O OH Aspartic Acid O H 2 N CHC H 2 C C O OH OH Cysteine O H 2 N CHC CH 2 SH OH Glutamic acid O H 2 N CHC H 2 C H 2 C C O OH OH Glutamine O H 2 N CHC CH 2 CH 2 C O NH 2 OH Histidine O H 2 N CHC CH 2 OH N NH Isoleucine O H 2 N CHC OH CH CH 3 H 2 C CH 3 Leucine O H 2 N CHC CH 2 OH CHCH 3 CH 3 Lysine O H 2 N CHC H 2 C H 2 C H 2 C H 2 C NH 2 OH Methionine O H 2 N CHC CH 2 CH 2 S CH 3 OH Phenylalanine O H 2 N CHC H 2 C OH Proline H N C O OH Serine O H 2 N CHC CH 2 OH OH Threonine O H 2 N CHC CH OH OH CH 3 Tryptophan O H 2 N CHC CH 2 OH HN Tyrosine O H 2 N CHC CH 2 OH Valine O H 2 N CHC OH CH CH 3 CH 3 OH

Primary Structure: Amino Acid Sequence

•20 amino acids • H 2 N R 1 O N H R 2 O H N O R 3 N H R 4 O H N R 5 O OH Average protein 300-400 amino acids = 30-45K Daltons Protein with 300 mers based on 20 amino acids: P =20 300 or 10 390 different possible sequences

Estimated

: 100,000 human proteins (coded by 30,000 genes)

Identified

: 10,000 human proteins

H 2 N H O N H H O H N H O N H H O H N H O HN H O H N H O N H H O H N H O OH Polyglycine H 2 N Me O N H Me O H N Me O N H Me O H N Me O HN Me O H N O Me N H Me O H N Me O OH Polyalanine HO H 3 N O N H O H N O N H O H N O O S Me meta-enkephalin

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 H

2

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

Large diameter egg Case fiber (Tubuliform) Flagelliform Aggregate Tubuliform Minor ampullate Major ampullate Acini form Pyriform Glue coating (Aggregate silk) (?) Capture Spiral (Flagelliform) 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 Fiber axis

Material

Dragline Silk Kevlar Rubber Nylon, type 6

Strength (GPa) Elasticity (%) Energy to break (J/kg)

1.1

3.6

0.001

0.07

35 5 600 200 4 x 10 5 3 x 10 4 8 x 10 4 6 x 10 4 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 Parachutes Fishing line Ballistics

Forced silking to obtain silk fibers

Spiders are anesthetized with CO 2 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) (Xaa = Tyr, Leu, Gln) and (GlyGlyXaa) n sequences - MW of MaSp1 ~ 275-320 kDa; Sp1+Sp2 ~ 700-750 kDa Repeating sequence of MaSp1 QGAG

AAAAAA

GGAGQGGYGGLGGQGAGQGGYGGLGGQGAGQGAG

AAAAAAA

GGAGQGGYGGLG GLGGYGGQGAGG

AAAAAA

GAGQGGRGAGQS SQGAGRGGLGGQGAG

AAAAAAA

GGAGQGGYGGLG GLGGYGGQGAGG

AAAAAA

GQGGRGAGQN SQGAGRGGLGGQAG

AAAAAA

GGAGQGGYGGLGGQGAGQGGYGGLG GLGGYGGQGAGG

AAAA

SAGAGQGAGQGGLGGQGAGG

AAAAAAA

GAGQGGLGGRGAGQS SQGAGRGGEGAG

AAAAAA

GGAGQGGYGGLGGQGAGQGGYGGLG GLGGYGGQGAGG

AAAAAA

GAGQGAGQGGLGGQGAGG

AAAA

GAGQGGLGGRGAGQS SQGAGRGGLGGQGAGAV

AAAA

GGAGQGGYGGLG GLGGYGRQGAGG

AAAAAA

GAGQGGRGAGQS NQGAGRGGLGGQGAG

AAAAAAA

GGAGQGGYGGLG GLGGYGGQGAGG

AAAAA

GQGGRGAGQN SQGAGRGGQGAG

AAAAAA

VGAGQEGIRGQGAGQGGYGGLG GAGGYGGQRVGG

AAAAAA

GAGQGAGQGGLGGQGAGG

AAAAAA

GAGQGGLGGRGSGQS SQGAGRGGQGAG

AAAAAA

GGAGQGGYGGLGGQGVGRGGLGGQGAG

AAAA

GGAGQGGYGGVG SSLRS

AAAAA

SAASAGS 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 (10 6 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 •Structural plant material •Cotton = >95% cellulose, wood = 50% per molecule •Stiff rods Conformation to the network.

•70% crystalline structure •T g = 227 ° C, T B = 298 ° C Tensile Strength: 800 MPa Modulus 75-100 GPa •MW of cellulose in 400,000 g/mol, corresponding to about 2200 D-glucose units •Well-organized water-insoluble fibers (20 nm diameter, 40,000 nm long) •The -OH groups form numerous intermolecular hydrogen bonds adding strength

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.

* OSO 3 D-glucuronic acid O O HO O O O HN O CH 3 HO N-acetyl-D-glucosamine O OH O OH O L-iduronic acid O O 3 S HN SO 3 O HO D-glucosamine OOC O OSO 3 O HO OSO 3 O NH SO 3 O D-glucosamine * Pentasaccharide unit of

heparin

antithrombin III.

responsible for binding to

Hyaluronic acid

D-glucuronic acid N-acetyl-D-glucosamine * HO O O O OH 1 HO O OH 3 2 O 1 O NH * O CH 3 Hyaluronic acid is the simplest acidic polysaccharide present in connective tissue MW of ~ 10 5 and 10 7 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.

Enzymes Enzymatic processing NaOH Dissolving Spinning acidic bath Fibre Other products Cellulose pulp Similar in both processes Waste Na 2 SO 4 / OH 2 O Alkalies CS 2 Alkali cellulose ksantogenation Cellulose xanthate Ripening Spinning acidic bath Fibre Other products

acetate rayon OH O HO O OH O glucose unit in cellulose fiber + 3 H 3 C O O O CH 3 acetic anhydride O H 3 C O O O O CH 3 O O O H 3 C O A fully acetylated glucose unit Tensile 10-250 Mpa Modulus 2 GPa Viscose rayon Cellulose OH NaOH Cellulose O Na + S C S S Cellulose O C S Na + Na + salt of a xanthate ester