ME406

Organization of Organisms: Kingdoms (Monera, Protista, Fungi, Plants, Animals). Bacteria are prokaryotic cells. Cyanobacteria: obtain energy through photosynthesis, monera domain, blue-green algae, releases oxygen, spurred life. Ciliates: protozoans with hair like organelles called cilia, common in water, protista domain. T4 virus: viruses are parasitic (infect host). Causes e.coli Ebola: single stranded RNA virus. Sea Urchin: Free reproduction, easy to obtain gamates, transparent eggs, Yeast: can reproduce but also break in half; fungi. Aspergillus: gene regulation, present in air. Drosophila Melanogaster (fruit fly), great for genetic studies, similar to humans, short life, easy to maintain, etc. C. Elegans: self consistency, highly regulated, transparent, can be frozen and revived Jellyfish: interesting healing and self defense. Octopus: intelligent, nervous system. Mollusks: hermissenda: good for neuroscience, easy to discriminate between species. 

Plant cells differ from Animal cells: chloroplasts (photosynthesis), cell wall, vacuole. Cytoskeleton: made up of microtubules, actin filaments, and intermediate filaments. Give cells shape + basis for movement and division.

Building Blocks of Cell: (Sugars –> Polysaccharides (unit of metabolism/energy), Fatty Acids –> FATs, LiPiDs (storing energy), Amino Acids –> Proteins (create structure, catalyze reactions), Nucleotides –> Nucleic Acids (source of energy). Monosaccharides condense to poly, Fatty Acids are components of cell membranes. Phospholipid: class of lipid that are major component of cell membranes (hydrophilic head, 2 hydrophobic fatty acid tails). ATP: energy carrier in our cells (type of nucleotide, made up of triphosphate, ribose, and adenine). DNA: 4 nucleotides (G/C, A/T…subtle differences in chemical structure/reaction). Building DNA: 1. phosphate + sugar = sugar phosphate 2. sugar phosphate + base = nucleotide 3. a lot of nucleotides = dna strand 4 2 strands join = double stranded dna, helix (double stranded is much more stable, C/G has three bonds, more stable than A/T which has two) uneven number: single stranded, maybe a virus. DNA Replication: genetic code: G, A, C, U….one gene = one protein, protein gives our body a specific trait for RNA. T turns to U. 

20 amino acids are the building blocks of proteins (4^3 = 64 possibilities, U, C, A, G are bases). 20,000 protein encoding genes in genome. polypeptides/proteins: polypeptides are chains of amino acids- work together to form protein. protein folding: Weak, non-covalent bonds (ionic, hydrogen (electrostatic attraction between 2 polar groups), van der waals forces (distance dependent) etc.) strength: ionic -> covalent -> Hydrogen -> van der waals. alpha helix: hydrogen bonds, stiff rigid, pitch 0.54nm, complete turn every 3.6 amino acids. beta sheet: pitch 0.7nm, repetition 2 nm. structure of coiled coil: takes two alpha helixes and coils them together (stronger). Occurs in nature. How protein engines might work: 1. enzyme binds to two substrate molecules and catalyzes a reaction 2. binding of substrate to enzyme rearranges electrons (reaction) 3. bending enzyme -> local strain -> favors a reaction. Allosteric Mechanism to make the cycle irreversable and walk in one direction. Chromomechanical Transduction (chemical energy -> mechanical work)

Methods: early microscopes (abbe, schott, zeiss) Leeuwenhoek- glass microscope 200x lens. Abbe: advancement for zeiss microscope. Resolution: 0.2 um, objective lens, condenser lens, resolution: (0.61*wavelength)/(n*sin(theta)) n = refractive index of medium, theta = half angular width of cone of rays collected by objective lens. Inverted microscope: light comes from above, good for micromanipulation. Transmission Electron Microscope (TEM): siemens 1964, digital/electronically controlled, chemical fixation (stabilizes specimens by killing them). Tissue sections for TEM (copper grid covered with carbon/plastic film) High Voltage (thicker speciman can be seen more easily, specimen heating reduced, resolution < 200 nm) Ultra High Voltage. Light diffraction limits resolution. Contrast within microscope methods: stained specimen. Fluorescence Microscopy the basic task is to let excitation light radiate the speciman and sort out the much weaker emitted light from the image. Quantum dots: nanoscopic semiconductor crystals as fluorescent probes. Confocal fluorescence microscopy: only taking that piece of information that is of interest (other light gets rejected) Cell Sorter: tissue is made up of different cells, after the cells are broken up, steam passes through the laser which separates the cells. Tissue Culture: grow individual cells. Chromatography: physical method of separation. Centrifuge purification of molecules: separates things by density. Faster you spin, more they separate. Chromatography: A. Ion exchange (covalently link molecules (+) and (-)) B. Gel-Filtration (size separation)

Polymerase chain reaction (application of DNA) purpose: to make a ton of DNA specific to one area. Allows you to amplify genes out of the dna of an organism. Detect DNA at scene of crime, change a gene. process: Heat double stranded DNA -> MELT (2 strands separate) -> anneal (bases go back together)…. 2^n # of dna molecules n = cycles. ensymatic DNA sequencing: modify the precursors (deoxy- allows extensions at 3′ end, dideoxy- prevents strand elongation at 3′ end by polymerase) electrophoresis: shortest lengths at bottom, useful for sequencing

Cytoskeleton: structure to help cells maintain shape/internal organization. Microtubules and subunits (OD = 24nm, ID = 12nm), is a hollow cylinder made of tubulin, alpha and beta stacked. Lumen is the space inside. 13 protofilaments in a microtubule. Flexible rigidity stronger as microtubule not protofilament. Actin filaments: linear polymer of globular actin subunits and occur as microfilaments in the cytoskeleton and as thin filaments which are part of the contractile apparatus in muscle and non muscle cells. (smaller and thinner than microtubules). D = 5-6nm. Nucleation and Elongation of Actin: g-actin particles come together and form nucleus (nucleation), subunits can assemble to either end (F-actin, elongation) -> the filament reaches steady state and as one g-acting particle adds on, one leagues from (-) side. Steady state: concentration of monomers. Time course of Actin Polymerisation: once you reach steady state there is no more growth. having enough available subunits is very important. Treadmilling: phenomenon observed in may ceulllular cytoskeletal filaments that occurs when one end of a filament grows in length while the other end shrinks resulting in a section of filament seemingly moving across a stratum or the cytosol. GTP –> GDP + Pi. Drugs disrupt microtubule dynamics. Fast Axonal Transport: precursor for how kinesin moves (molecular organization of kinesin: heavy chain, coiled coil, light chain). Kinesin “walks” along and moves vesicle to (+) end. in vivo = inside, in vitro = outside. no matter how many kinesin are attached to a microtubule, it will move at the same speed. more ATP = faster microtubule moves. Laser Trapping: shine a beam-force generated from beam, if a lot of light gets through, detect location. Use this to determine how much force is being generated and how far the kinesin moves. Myosin: is to actin, what kinesin is to microtubules. NATURE HOWARD ET AL: do biological motors move with regular steps? Kinesin moves with 8-nm steps (one dimer of microtubulin, so it walks from tubulin to tubulin) procedure: deposit silica beads carrying single molecules of motor protein kinesin on microtubules. if microtubule is bound to glass then kinesin moves. a single kinesin can move a microtubule (keep diluting the solution). Muscles: sarcomere structure (actin = thin, myosin = thick, myosin is moving) I A I bands, I bands are less dense = actin + myosin, A band = just actin. sliding filament model relax, contracted, 2.2um fundamental. Neuromuscular junction (nuerons -> muscles) cells have electric potential. Nerve impulse –> acetylcholine out, na ion flow in, calcium channel opens, let calcium out -> causes muscle Myofibril: collection of mini fibers Sarcoplasmic reticulum: holds calcium Electromechanical Coupling: 20-30ms, reaction time of body. Twitches/Tetanus: holding something, staying contracted = tetanus. 1 twitch = 1 release of calcium. 

DNA: packed into chromosomes in cells. Genome size = 10^5 – 10^12 = # of base pairs in genome. Of genes, only 1% codes DNA. Easily subject to radiation damage. Persistence Length of Polymers: length at which angles aren’t related anymore (DNA has a short persistence length) quantifies stiffness. Microtubule has longer persistence length. 100 L = very stiff. DNA Molecule: 3′ end: hydroxyl 5′ end: phosphate, pitch: 0.34nm per base pair = 3.4A. DNA encodes proteins which give cells characteristic properties. DNA serves as its own template for duplication. (Unzip dna into 2 strands, RNA walks along DNA and adds that piece, DNA Polymerase (leading strand 5 to 3, lagging strand 3 to 5). Okazaki fragments formed on lagging strand. X & Y are gender (chromosomes), 23 pairs. Mitosis: separation; reproduction, how chromosomes are separated. Telomeres can predict life expectation. 2* # of chromosomes = 46 replication origins to copy human genome. Anaphase (shorten) Interphase (period after separation and before) Chromatin packing (dna wraps around histone)