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Living Parts •Prokaryotes, Eukaryotes •Tissue – groups of cells together for certain specialized functions, differentiated cells •Tissue – 14 major types of tissues in animals epithelial, connective, nervous, muscle, etc. http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookAnimalTS.html •Cell – over 200 types in a vertebrate Long – eg. nerve cells Some do not divide for ~ 100 years Some divide rapidly, ~ few hours Components of a Cell (Eukaryotes) ~70% water 4% small molecules 15-20% proteins 2-7% DNA/RNA 4-7% membrane Picture from on-line biology book, http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookCELL2.html Membrane • Lipid bi-layer Phospholipids and other lipids hydrophilic, hydrophobic • Small molecules and membrane-bond proteins • Semi-permeable / Osmosis N2, O2, water, glycerol, glucose, sucrose, Ions, etc. http://en.wikipedia.org/wiki/Cell_membrane Picture from : http://www.cbc.umn.edu/~mwd/cell_www/chapter2/membrane.html Cytoplasm • Cytoskeleton – fibrous protein complexes maintain shape, anchoring, moving actin filaments microtubules • Ribosome – protein synthesis • Mitochondrion – energy • Endoplasmic reticulum (ER) – mesh of membrane, protein synthesis and transport • Lysosomes, Golgi, vesicles etc. • A good reference site http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookCELL2.html Nucleus • Nuclear membrane • Nuclear envelope with pores • DNA/RNA and some proteins • A good reference site http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookCELL2.html Nucleic Acids • DNA – polymers of deoxyribonucleic acids, ds • Nucleotide: 3 components: base (purine/pyrimidine) sugar (ribose/deoxyribose) phosphate group C (cytosine) T (thymine, DNA) U (uracil, RNA) RNA: in both nucleus and cytoplasm, ss 3 types: mRNA, rRNA and tRNA • Picture from on-line biology book • http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookCHEM2.html A (adenine) G (guanine) Protein-a chemical view • A chain of amino acids folded in 3D • Peptide Protein backbone N / C terminal • Picture from on-line biology book Amino Acids • 20 types in nature • Different properties – side chain Generally: • Positively charged – Arg, His, Lys • Negatively charged – Asp, Glu • Polar but uncharged – Ser, Thr (OH), Asn, Gln(CO) • Special – Cys, Gly, Pro • Hydrophobic – Ala, IIe, Leu, Met, Phe, Trp, Tyr, Val, • A good reference site http://www.emc.maricopa.edu/faculty/farabee/BIOBK/BioBookCELL2.html Protein – a 3D view • Bond length, bond angle – fairly restricted • Torsion angles on backbone – (phi), (psi), (omega) • , mostly plane(180°, rare case 10°in cis) • , , free but with an average characteristic distribution- Ramachandran plot • Picture from http://www.expasy.org/swissmod/course/text/chapter1.htm Torsion Angles • Dihedral angles (phi), (psi), (omega) N C C N Secondary structures • Helix - hydrogen bond (CO)i-(NH)i+4 -helix (3.613) 1.5Å / residue • -sheet is composed of multiple -strands •Hydrogen bond between two -strands • Zig-zag backbone, side-chains opposite directions , ~30°/residue twist, mostly antiparallel • Turn, loop/coil Picture from www.expasy.org site http://www.expasy.org/swissmod/course/text/chapter1.htm Protein tertiary and quaternary structure • Tertiary – 3D folding of a polypeptide chain involves non-local interaction • Quaternary – multiple chains/multi subunits PDB: http://www.pdb.org SCOP database – protein classification From DNA to Protein • Genome, genes, chromosome, proteome • Overview of HGP • Transcription (DNA-mRNA) • Translation (mRNA-polypeptide) - Gene expression Picture from doegenomics.org http://www.ornl.gov/T echResources/ Human_Genome/proj ect/info.html Transcription • Initiation, Elongation and Termination • Central enzyme: RNA polymerase Picture from http://edtech.clas.pdx.edu/gene _expression_tutorial/transcriptio n.html • RNA polymerase bind to promoter site, e.g. in bacteria 35 BP upstream of start: RNA polymerase binding site (TTGACA) 10 BP upstream of start: box (TATAAT) - sigma factor site •Promoter sequence determines transcription level Transcription in Eukaryotes • More complicated process • RNA Splicing – intron and exon Picture from http://www.intouchlive.com/home/frames.htm?http://w ww.intouchlive.com/cancergenetics/genefx.htm&3 • Alternative splicing – diversity of proteins Translation • Genetic coding • What is a codon? •Ribosome bind upstream region of mRNA •tRNA bind to specific amino acid (AUG) on mRNA to start •tRNA brings a.a. to ribosome •At least one tRNA exists for each amino acid Example of a tRNA http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/T/Translation.html Picture from http://edtech.clas.pdx.edu/gene _expression_tutorial/translation. html Regulation in gene expression • Various needs for gene expression • Spatially and timely different steps in eukaryotes • Prokaryote – e.g. lac gene regulation http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/L/LacOperon.html • Eukaryotes Altering rate of transcription Rate of transcript processing, stability of mRNA, efficiency of ribosome • Basel promoter, upstream promoter • Enhancer, silencer • Transcription factors http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/P/Promoter.html Experimental techniques • Identify size of protein/DNA e.g. gel electrophoreses • Identify proteins e.g. using antibodies - structural • Sequencing peptide e.g. mass spectrometry • Sequencing DNA/RNA • Determine some 3D protein structure • Molecular cloning, producing large amount of genes and proteins Recombinant DNA technology • Restriction enzyme, ligase • Vector – plasmid, bacteriophage (virus) Recombinant DNA technology • Cleave DNA • Vector to carry DNA for cloning • Transform bacteria • Grow bacteria • Screen for cloned DNA • Revolutionized biology • An example • http://www.biology.arizona.edu/molecul ar_bio/problem_sets/Recombinant_DN A_Technology/05t.html Related techniques • cDNA, vs. genomic DNA reverse transcriptase represent currently active mRNA population function, stage of the cell A cool animation http://www.maxanim.com/genetics/cDNA/cDNA.htm • Polymerase Chain Reaction (PCR) in-vitro amplification of a region of DNA with known sequence primer, template DNA polymerase • http://en.wikipedia.org/wiki/Polymerase_ch ain_reaction Protein Structure Determination • X-ray crystallography soluble, medium size, some viruses usually difficult for large proteins • Nuclear Magnetic Resonance (NMR) small , multi-dimensional NMR • Other developing methods e.g. electron cryomicroscopy • Structural genomics X-ray crystallography X-ray Protein Crystals Diffraction data Phase Structure Electron density map Sequence Grow suitable crystals – tricky Solving structure – mostly a mature technique Electron cryo-microscopy • 2D crystallography – e.g. membrane proteins • Non-crystalline – e.g. viruses, large complexes, helical objects • Take 2D images using TEM • Computationally build 3D structure • Computationally more intensive http://en.wikipedia.org/wiki/Cryo-electron_microscopy