Synthetic Polymers vs. ProteinsProtein Adsorption on Biomaterial SurfacesModels for Protein Adsorption3.051J/20.340J 1Lecture 5: Protein-Surface Interactions Importance of Protein-Surface Interactions: • Modulate cell adhesion • Trigger the biological cascade resulting in foreign body response • Central to diagnostic assay/sensor device design & performance • Initiate other bioadhesion: e.g., marine fouling, bacterial adhesion Fundamentals on Proteins: • Largest organic component of cells (~18 wt% /H2O =70%); extra-cellular matrix, and plasma (7wt% /H2O=90%). • Many thousands exist—each encoded from a gene in DNA. • Involved in all work of cells: ex, adhesion, migration, secretion, differentiation, proliferation and apoptosis (death). • May be soluble or insoluble in body fluids. Insoluble proteins—structural & motility functions; can also mediate cell function (ex., via adhesion peptides) Soluble proteins—strongly control cell function via binding, adsorption, etc. • Occur in wide range of molecular weights.3.051J/20.340J 2“Peptides” (several amino acids): hormones, pharmacological reagents e.g., oxytocin: stimulates uterine contractions (9 a.a.) aspartame: NutraSweet (2 a.a.) “Polypeptides” (~10-100 amino acids): hormones, growth factors e.g., insulin: 2 polypeptide chains (30 & 21 a.a.) epidermal growth factor (45 a.a.) “Proteins” 100’s-1000’s of amino acids e.g., serum albumin (550 a.a.) apolipoprotein B: cholesterol transport agent (4536 a.a.) Protein Functions: • Structural/scaffold: components of the extracellular matrix (ECM) that physically supports cells e.g., collagen—fibrillar, imparts strength; elastin—elasticity to ligaments; adhesion proteins: fibronectin, laminin, vitronectin—glycoproteins that mediate cell attachment (bonded to GAGs) • Enzymes: catalyze rxns by lowering Ea thru stabilized transition state, via release of binding energy e.g., urease—catalyzes hydrolysis of urea3.051J/20.340J 3 • Transport: bind and deliver specific molecules to organs or across cell membrane e.g., hemoglobin carries bound O2 to tissues; serum albumin transports fatty acids • Motile: provide mechanism for cell motion e.g., via (de)polymerization & contraction e.g., actin, myosin in muscle • Defense: proteins integral to the immune response and coagulation mechanism e.g., immunoglobulins (antibodies)—Y-shaped proteins that bind to antigens (foreign proteins) inducing aggregate formation fibrinogen & thrombin—induce clots by platelet receptor binding • Regulatory: cytokines—regulate cell activities e.g., hormones: insulin (regulates sugar metabolism); growth factors3.051J/20.340J 4Protein Structure Proteins have multiple structural levels… 1. Primary Structure ¾ comprised of amino acid residues: - -CHR- - OHCN¾ peptide (amide) bond CONH is effectively rigid & planar (partial double-bond character) ¾ directional character to bonding: amino acids are L stereoisomers [after A. L. Lehninger, D. L. Nelson and M. M. Cox. Principles of Biochemistry, pg. 171.] [after A. L. Lehninger, D. L. Nelson and M. M. Cox, Principles of Biochemistry, pg. 115.] Figure by MIT OCW.Figure by MIT OCW.3.051J/20.340J 5¾ side groups R have variable character [after A. L. Lehninger, D. L. Nelson and M. M. Cox. Principles of Biochemistry.] Figure by MIT OCW.3.051J/20.340J 62. Secondary Structure Spatial configuration determined by the rotation angles ϕi & ψi about the single bonds of the α-carbons (ϕi ,ψi) are independent of (ϕi+1,ψi+1) [after P. J. Flory. Statistical Mechanics of Chain Molecules, pg. 251.] Ramachandran plots: designate permitted ranges of ϕ & ψ for a.a. residues Image removed due to copyright considerations[from A.L. Lehninger, D.L. Nelson & M.M. Cox, pg. 171.] Figure by MIT OCW.3.051J/20.340J 7 β−sheets ¾ backbone has extended “zigzag” structure ¾ stabilized by intermolecular H-bonding between –NH and C=O of adjacent chains α−helices ¾ stabilized by intramolecular H-bonding between C=O of residue i and –NH of residue i+3 (requires all L or D stereoisomers) [after A. L. Lehninger, D. L. Nelson and M. M. Cox. Principles of Biochemistry, pg. 169.] [after P. J. Flory. Statistical Mechanics of Chain Molecules, pg. 287]Figure by MIT OCW.Figure by MIT OCW.3.051J/20.340J 8 ¾ natural abundance  most common secondary structure in proteins  in fibrous proteins: α−keratins (hair, skin,…)  in globular proteins: avg. ~25% α−helix content 3. Tertiary & Quaternary Structure ¾ Tertiary: folded arrangements of secondary structure units ¾ Quaternary: arrangements of tertiary (polypeptide) units Example: hemoglobin [from A.L. Lehninger, D.L. Nelson & M.M. Cox, pg. 187.] Image removed due to copyright considerations3.051J/20.340J 9Synthetic Polymers vs. Proteins Property Synthetic Polymers Polypeptides Molecular Wt. 1000-106 g/mol 1000-106 g/mol ( typ. <2000 a.a.) Molecular Wt. Distribution Always > 1 (Mw/Mn) Always ≡1 Sequence i. 1-3 types of repeat units ii. many chemistries i. many side groups ii. always amides Solution Structure Random coils or self-avoiding random coils Rg~N0.5 (θ solvent) Rg~N0.6 (good solvent) Globular –“condensed” chains (ρ~1.36 g/cm3) (hydrophobic R groups sheltered from H2O) Rg~Naa0.33 Available Conformations Ωran ∼ zN (z = # n.n.) ΩSA ∼ z’N Ν1/6 << ΩranΩ ~1 (can ↑ if bound or adsorbed!) Secondary Interactions van der Waals, H-bonds, electrostatic, “hydrophobic effect” Same as synthetic, with“ lock-and-key” topology Polypeptides can transform to “random coil” conformations, through: ¾ changes in temperature ¾ changes in soln. pH or composition (e.g., added salts, urea) ¾ adsorption to surfaces ⇒ changes physiological function!3.051J/20.340J 10Protein Adsorption on Biomaterial Surfaces Background a) Protein activity varies in adsorbed vs. solvated state Why? 1. higher local concentration— function may be conc. dependent e.g., cell adhesion increases with adhesion peptide concentration 2. change in reactivity—access to “active” a.a. sequence ↑ or ↓ ⇒ enhanced or reduced binding capability e.g., fibrinogen: platelets