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Bionanotechnology Dr Cait MacPhee ([email protected]) Dr Paul Barker ([email protected]) Mondays 12 pm, Tuesdays 11 am Syllabus The molecules of life Proteins (6 lectures) background as components in nanodevices biomolecular electronic devices electron transport and photosynthesis as fibrous materials in motion – molecular motors DNA (3 lectures) background as components in nanodevices: part I as components in nanodevices: part II Lipids (1 lecture) background; as components in nanostructures: artificial cells (liposomes and membrane nanotubes) Bio-inorganic composites (1 lecture) composites – including butterfly wings, diatoms, mineralisation The whole cell Cell mechanotransduction (1 lecture) bringing together physical, life, and applied sciences; bone cell mechanobiology Cell motility (1 lecture) how cells travel and navigate through 2and 3 dimensional environments Biomaterials (1 lecture) surface science/ surface chemistry; tissue engineering Nanomedicine (1 lecture) Nanotherapeutics, real and imagined · Qdots and developmental biology Ethical considerations (1 lecture) risk/benefit analysis focusing on bionanotechnology Suggested texts: Nanobiotechnology, edited by CM Niemeyer and CA Mirkin Bionanotechnology, DS Goodsell http://bionano.rutgers.edu/mru.html Proteins The basics • Proteins are linear heteropolymers: one or more polypeptide chains • Repeat units: one of 20 amino acid residues • Range from a few 10s-1000s • Three-dimensional shapes (“folds”) adopted vary enormously – Experimental methods: X-ray crystallography, electron microscopy and NMR (nuclear magnetic resonance) L-amino acids The peptide bond • has partial (40%) double bond character • ~ 1.33 Å long - shorter than a single, but longer than a double bond • Ca usually trans • the 6 atoms of the peptide bond are always planar • N partially positive; O partially negative, gives rise to a significant dipole moment d- Ca d+ Ca Free backbone rotation occurs only about the bonds to the a-carbon c Y F Y: rotation about the Ca-C bond F rotation about the Ca-N bond Steric considerations restrict the possible values of Y and F Ramachandran plots Antiparallel b-sheet Parallel b-sheet Triple coiled-coil a-helix (L) a-helix (R) Flat ribbon Used to display which conformations are allowed. All the disallowed conformations are sterically impossible because atoms in the backbone and/or side chains would overlap. The amino acids isoleucine tryptophan asparagine glutamate alanine The amino acids • Hydrophobic: Alanine(A), Valine(V), phenylalanine (Y), Proline (P), Methionine (M), isoleucine (I), and Leucine(L) • Charged: Aspartic acid (D), Glutamic Acid (E), Lysine (K), Arginine (R) • Polar: Serine (S), Theronine (T), Tyrosine (Y); Histidine (H), Cysteine (C), Asparagine (N), Glutamine (Q), Tryptophan (W) The disulphide bond • Only in extracellular proteins • Formed by oxidation of the SH (thiol) group in cysteine amino acids • Forms a covalent cross-link between the Sg atoms of two cysteines Protein structure Hierarchy of structures 1° Sequence 2° a/b 3° Packaging 4° Assembly Hierarchy of structures Primary structure: sequence of amino acids Secondary structure: • Alpha helix • Beta sheet • Beta turns Local structures stabilized by hydrogen bonds within the backbone of the chain The a-helix C • One of the two most common elements of secondary structure • Right-handed helix stabilized by hydrogen bonds • amide carbonyl group of residue i is Hbonded to amide nitrogen of residue i+4 • 3.6 amino acids per turn • acts as a strong dipole • H-bonds are parallel to the axis of the helix • Y = -47, F = -57° N The a-helix C • One of the most closelypacked arrangements of amino acids • Sidechains project outwards • Can be amphipathic • Average length: 10 amino acids, or 3 turns • Varies from 5 to 40 amino acids N The coiled-coil • “Supersecondary” structural motif • Two or more a-helices wrapped around each other • Stable, energetically favorable protein structure • “Heptad Repeat”: pattern of side chain interactions between helices is repeated every 7 Amino Acids (or every two “turns”) The coiled-coil • Heptad repeat in sequence – [a b c d e f g]n • Hydrophobic residues at “a” and “d” • Charged residues at “e” and “g” +/b e a Hydrophobic residues at “a” and “d” f d c g Charged residues at “e” and “g” The coiled-coil C C Residues at “d” and “a” form hydrophobic core Residues at “e” and “g” form ion pairs -/+ b g e a +/- c d f f a d c g e +/- b -/+ N N The b-Pleated Sheet • Composed of bstrands, where adjacent strands may be parallel, antiparallel, or mixed • Brings together distal sections of the 1-D sequence • Can be amphipathic The b-Sheet Mixed Parallel AntiParallel Loops • Regions between a helices and b sheets • Various lengths and three-dimensional configurations • Located on surface of the structure (charged and polar groups) • Hairpin loops: complete turn in the polypeptide chain, (antiparallel b sheets) 2 3 1 • Highly variable in sequence 4 • Often flexible • Frequently a component of active sites Amino acid propensities Helix Sheet Ala High inhibitory Cys inhibitory Intermediate Asp inhibitory Breaker Glu High Breaker Phe Intermediate Intermediate Gly Breaker No preference His No preference Intermediate Ile Intermediate High Lys Intermediate No preference Leu High Intermediate Met High Intermediate Asn No preference No preference Pro Breaker Breaker Gln Intermediate Intermediate Arg inhibitory inhibitory Ser inhibitory No preference Thr inhibitory Intermediate Val Intermediate High Trp Intermediate Intermediate Tyr No preference High Driving forces in protein folding • Stabilisation by formation of hydrogen bonds • Burying hydrophobic amino acids (with aliphatic and aromatic side-chains) • Exposing hydrophilic amino acids (with charged and polar side-chains) • For small proteins (usually > 75 residues) – Formation of disulfide bridges – Interactions with metal ions Hierarchical organisation Tertiary structure • Packing of secondary structure elements into a compact independently-folding spatial unit (a domain) • Each domain is usually associated with a function (“Lego”) • Comprises normally only one protein chain: rare examples involving 2 chains are known. • Domains can be shared between different proteins. Ig EG EG EG Ig F3 Ser/Thr Kinase Quaternary structure • Assembly of homo- or heteromeric chains • Symmetry constraints Hierarchy of structures 1° Sequence 2° a/b 3° Packaging 4° Assembly Protein folds • ~70,000 proteins in humans • ~21,000 structures known • Only 6 classes of protein folds – Class a: bundles of a helices connected by loops on surface of proteins – Class b: antiparallel b sheets, usually two sheets in close contact forming sandwich – Class a/b: mainly parallel b sheets with intervening a helices; may also have mixed b sheets (metabolic enzymes) – Class a+ b: mainly segregated a helices and antiparallel b sheets – Multidomain proteins(a and b) - more than one of the above four domains – Membrane and cell-surface proteins and peptides excluding proteins of the immune system Prosthetic groups Small blue proteins (azurin) His R Glu N Cu Cys His Cys His S Cu O Cu S Met Cys His Cytochrome c oxidase Retinal C + N R Haemoglobin