Cell bilogy BY330
Cell bilogy BY330 330
Popular in Cell Biology
Popular in Biology
This 41 page Bundle was uploaded by Blake Bearden on Monday January 25, 2016. The Bundle belongs to 330 at University of Alabama at Birmingham taught by Mickie Lynn Powell in Fall 2015. Since its upload, it has received 26 views. For similar materials see Cell Biology in Biology at University of Alabama at Birmingham.
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Date Created: 01/25/16
History Hooke O coined the term cell 0 observed dead plant cell from a piece of cork 0 used a primitive microscope Leewenhoek 0 observed pond water 0 1st person to identify protozoan and bacteria Scleiden botanist amp Schwann zoologist 0 Cell Theory 1 all organisms consist of 1 cells 2 cell is the basic unit of life 3 all cells arise from preexisting cells 0 In 1800s Higher Resolution Microscopes 0 Resolution the ability to distinctly tell the difference between 2 points 0 Know measurements and how to do conversions I m gt dm gt cm gt mm gt micrometers gt nm gt A I 0 What limits cell size I Surface Area of membrane and Volume I SA squared V cubed I we want large amount of SA and small volume for maximum gas and nutrient exchange What is a Cell 0 smallest unit of life 0 makes up all living organisms 0 take on specialized tasks in multicellular organisms Structure of Cells 0 membrane bound structure that contains 0 biomolecules 0 nucleic acids RNA and DNA 0 proteins catalystsenzymes 0 lipids 0 polysaccharidessugars Carbs Cell Types 1 Prokaryotic O Mycoplasms genus of bacteria that lack a cell wall and are hard to kill 2 Eukaryotic 0 membrane bound organelles O ones that become specialized into multicellular organisms 3 Side Note Viruses are not organisms they have to use other organisms machinerycannot live by themselves Components of cells 0 Chemical components 0 Small molecules100s of Da 0 assembled into larger Macromolecules200000Da or more Chemical Components of Cells Chonps 90 0 Carbon Hydrogen 0 Oxygen Nitrogen Phosphorus in DNA and RNA Sulfer Characteristics of Water Not organicquot makes up 70 of cells by weight Universal Solvent 0 water hold on the heat very well high BP and MP Hight Surface Tension 2 Covalent Bonds holds water together mutual sharing 0 O has partial negative charge 0 H have partial positive charge 0 Yielding a polar molecule 0 Hydrogen bond hold different water molecules together KNOW HOW TO DRAW HYDROGEN BOND I make sure you draw dipole moments I 0 Relative Inert Freezes and Floats Interactions with Water 0 Hydrophilic water loving favorable interactions with water I NaCl separate and get surrounded by water molecules I O Hydrophobic phobia of water do not have favorable interactions of water I Methyl groups do not get surrounded by water I formsfolding of molecules 0 Amphipathic having hydrophobic and hydrophilic regions Carbon ElementAtom Make living molecules Si is another molecule similar 0 Small atomic 6 0 4 outer shell electrons valence of 4 potential to form 4 covalent bonds 0 Can form large molecules With no upper size limit Functional Groups found in carbon molecules gives them specificity CH3 0 COOH reactive OH reactive NH2 Small Molecules 0 Carbon compounds 1001000Da usually free in solution 0 for pool of intermediates to make macromolecules 0 nearly 1000 different kinds grouped by common characteristics 0 Examples 1 Amino Acids monomer gt Proteinspolymer I Structure I alpha carbon I amino group I carboxyl group I R groups I hydrophobic assist in protein folding I hydrophilic I acidic I basic I l Functions of AA 1 Energy 2 Building Blocks for Proteins 2 Simple Sugars monomer gt Carbohydrates polymer I monosaccharides CH20n n37 I example Glucose C6H1206 and Deoxyribose I Know structure of Glucose I I ketoses and aldoses react differently I disacchardies 2 sugars combined I example Sucrose dehydration reaction I Function of Sugars I Oxidized for Energy I Structure of plants Cellulose I Cellular Recognition Receptors 3 Fatty Acids gt Lipids operational term doesn t have monomer not soluble in polar solutions examples I fatty acids triglycerides phospholipids I steroids cholesterol Structure long chain of carbons that ends with a carboxyl group Saturated Fat no double bonds in the FA tend to be Solid butter Unsaturated Fat one or more double bonds in the FA tend to be Liquid olive oil I If DB is on C3 Omega 3 FA If DB is on 06 Omega 6 FA I I Functions of Fatty Acids 1 Energy fats have twice the amount of energy stored than any other molecule high energylow weight I Triglycerides whats stored in your fat cell cleave off triglyceride when you burn energy Hydrophobic I 2 Structure ex phospholipids of cell membrane I I Organic groups attached to phosphate group Hydrophilic portion I choline I ethanol I inostitiol I Phospholipid has hydrophobic tails and hydrophilic heads0rganic head gtfatty acid I 3 Cell Signaling 4 NucleotidesNucleosidesmonomer gt DNARNApolymer I Nucleotide sugar base phosphate I Structure I I Function of Nucleotide I preserve information in biological systems I carrier of energy ATP I signaling molecule cAMP I Nucleoside sugar base gt No side I Components I Sugar I Ribose RNA I Deoxyribose DNA I I Macromolecules 0 10000 1000000 Daltons 0 Assembled from low mw subunits Chains formed by covalent bonds 0 Shared electrons 0 Strong 0 Form backbone Function with weaker bonds ie hydrogen bonds 0 Non covalent bonds I NaCl ionic compound I NaCl with water gtgtgt I Hydrogen bonds I I Van der Wals forces I Weak electrostatic attractions Enzyme function is through weak bonds Information and actiVity is expressed through Weak bonds 0 binding specificity 0 Stabilize macromolecules associations so they can act as building blocks Peptide bond strong covalent bond Dehydration reaction A Polypeptide long chain of amino acids R groups will want to reach lowest energy state ie hydrophobic and hydrophilic molecules will fold so that they are where they want to be 0 Chaperones will assit in folding Can shield other molecules and may protect proteins during times of shock ie heat shock proteins Globular and fibrous proteins ie actin 0 Not all conformations are functional How many unique 300 amino acid polypeptides can you make 20quot300 unique polypeptides possible Estimated 80000 functional proteins because not all 3d structures are functional Functional 3D protein structure 0 Stable confirmation low energy 0 Flexible need to be able to interact with other structures 0 Single amino acid change destroys function 0 Primary structure 0 long chain of amino acids 0 Secondary structure 0 Interactions of R groups with other R groups and environment 0 Alpha helix 0 Beta sheet Tertiary structure 0 Interaction of secondary structures in a single polypeptide chain 0 Quaternary structure 0 Interaction of two or more polypeptide chains 0 Dimer dyemer 2 polypeptide chains I Homodimer both polypeptide chains are the same I Heterodimer two different chains 0 Trimer 3 pp chains 0 Polymer multiple pp chains I Actin filaments I Tubes made of tubulin I Viral coats Protein functions Initiation factors Elongation factors 0 Structural fibril proteins ie exoskeletons Enzymes most important Ligand substance that binds to protein 0 Ion 0 Small molecule 0 Another macromolecule protein DNA RNA 0 Binding is selective and high affinity Ligands bind through weak bonds Induced fit doesn t have to exactly match binding site but will conform to binding site or binding site will conform to ligand Allosteric proteins have two or more binding sites Often regulatory sites Proteins Binding sites ligands binds to binding site Domains 3D structure of folded polypeptide Enzymes 0 class of proteins that act as a catalyst 0 enhance favorable reactions enzyme ligands can be regulatory or substrates 0 enzyme substrate gt product enzyme enzymes allow bonds to break covalent bonds in a controlled way to convert substrate to product 0 enzymes are not consumed in the reaction 0 ES gt ES gtEP gt EP Rate of Product Formed 0 amount on enzyme 0 amount of substrate affinity of substrate binding site High affinity enzyme decrease Km Low affinity enzyme increase Km Competitive Inhibitor if you increase substrate it will overcome a competitive inhibitor higher probability Km changes 0 pharmeceutically O Noncomeptitve inhibitor only the Vmax changes 0 Bioenergetics Laws of Thermodynamics 0 energy can be neither created not destroyed 0 all systems tend to go to a state of maximum entropy disorder 0 Energy 0 Released through the oxidation of organic molecules I Oxidation transfer of electrons from one atom to another I Reduction addition of electrons I LEO goes GER I Oxidation is the most favorable state lowest energy 0 Gibbs free energy Coupled reaction Use energy from a favorable reaction to run reactions that are not favorable Energy storage molecules I Readily transferable energy group I High energy electrons I Coenzymes may be required Requirements for biosynthesis ATP gtgt 1113 kCalmol 0 Reducing power 0 H and 2 electrons hydride ion 0 Release of glucose energy needs to be controlled and slow 0 Metabolism 0 Sum of all chemical processes carried out in the cell 0 Catabolic reaction breaking large molecules down into smaller molecules 0 Usually exergonic deltaG Anabolic reactions small molecules into larger molecules 0 Usually endergonic deltaG Autotrophs 0 Obtain energy from light or inorganic compounds Heterotrophs 0 Obtain energy by consuming and breaking down organic molecules Glucose 0 preferred energy source Aerobic in the presence of oxygen Anaerobic in the absence of oxygen 0 One molecule of glucose releases 720 kCal 0 Only 13 gets captured as ATP 0 23 lost as heat Which helps maintain core body temp Glycolysis Anaerobic occurs in Cytoplasm O deltaG44kCal just from glycolysis 1 ATP 73 kCal 27344 X 100 30 glycolysis is only about 30 efficient Substrate level phosphorylation O O O 2 Phases 1 Energy Investment Phase 2 ATP 0 Between glucose and glucose6phosphate I Enzyme used Hexokinase kinase gt move phosphate I Key Regulatory Point 0 Between fructose6phosphate and fructose 16bisphosphate I Enzyme used PFKl Phosphofructosekinasel KEY regulatory enzyme for glycolysis 2 Energy Generating Phase 4 ATP produced via substrate level phosphorylation 0 2 glyceraldehyde3phosphate are converted to 2 pyruvate molecules 2 ATP produced each I Enzyme used Pyruvate Kinase 3 Net ATP Produced 42 2 ATP 0 If asked on exam to determine how many ATP are generated only answer the question asked I Example If starting molecule are 2 G3P and end product is pyruvate how many ATP will be produced 4 ATP I Example if you start at a molecule of glucose gt pyruvate what is the net ATP produced 2 ATP Electrons 0 Lose electrons through the entire pathway of metabolism to NAD gt NADH 2 electrons an H are transferred 0 NADH carries 2 e 0 Glycolysis yields Net 2 NADH H Fermentation regenerate electron acceptors in the form of NAD Key Regulatory Points associated with deltaG free energy steps more favorable 1 Hexokinase synthesis of sugars for DNA and RNA 2 PFKl speeds up or slows down reactions Big regulating point 0 has 2 binding sites on PFKl I one binds to ATP Slows down I one binds to fructose 26BP speeding it up I f6P gt f16BP I to increase reaction rate we need to decrease Km I f6P PFK2 gt fructose 26BPbinds to PFKl decreasing km increasing rate of reaction 3 Pyruvate kinase To run Glycolysis in Reverse to get back up deltaG use different enzymes to make it favorable again 1 pyruvate carboxylase and PEP instead of pyruvate kinase 2 fructose 16bisphosphatase instead of PFKl 3 glucose 6phosphatase instead of hexokinase Enzyme Inhibitor Activator Hexokinase ATP Phosphofructokinase 1 ATP ADP AMP Fructose26bisphosphate fructose6phosphate Pyruvate kinase ATP fructo 16bisphosphate For Exam 0 Gluconeogenesis 0 know 4 enzymes that are unique to gluconeo 0 you do not need to know intermediates 0 Glycolysis 0 know the intermediates 0 know the 3 key regulatory points and what controls them 0 energy in and out ATP electrons NADH H Pyruvate Dehydrogenase Complex Pyruvate needs to be transported from cytosol to mitochondria 0 occurs in the intemembrane space 0 Once pyruvate is transported 0 Pyruvate Dehydrogenase Complex Enzyme I a C02 come off of the pyruvate I oxidation of 1C molecule 1 C02 per pyruvate 2 CO2 I NAD converted into NADH generates 2 NADH I Coenzyme A comes in and attached to 2 carbon molecule forming acetyl coenzyme A I Kreb Cycle TCA cycle 9 Steps 0 Complete oxidation of 2C molecules 0 have to do this twice because you have 2 molecules of pyruvate 0 begin and ends With same substrate 0 oxaloacetate is regenerated 0 Net products from 1acety1 CoA 0 ATP equivalent GTP 0 1 FADH2 carries 2e 0 3 NADH H 6e 0 2 CO Regulation of Kreb Cycle in mitochondrial matrix 0 substrate availability 0 product inhibition 0 Regulatory enzymes 0 Citrate Synthase O Isocitrate dehydrogenase O alphaketogluterate DH Complex 0 downregulate WATP O upregulate WADP Ca2 Kreb Cycle Video 2 C02 come out and then oxaloacetate is being reformed Oxidative Phosphorylation chemiosmotic process mitochondria has 2 phospholipid bilayers 0 the inner phospholipid bilayer has increased SA due to cristae I Where the Electron Transport Chain is ETC Electron Transport inner membrane of mitochondria Oxidative Phosphorylation Lets look at What all we have and Where these molecules are located Glycolysis in cytosol 0 Net 2 ATP Gross 4 ATP 2 NADH H 4e PDH 2x in mitochondria 2 NADH H 4e 0 2 C02 Krebs 2x in mitochondria 6 NADH H 12e 2 FADH H 4e 0 2 ATP equivalents GTP 4 CO2 Total there are 12 molecules carrying electrons that need to be oxidized ETC does the job NADH Dehydrogenase Complex Complex 1 on inner membrane of mitochondria 0 about 40 polypeptide chains in size 0 accepts electrons from NADH by oxidation 0 e passed through 7 irons sulfur center complexes Complex II FADH2 0 Complex 2 only carries electrons from FADH2 the electrons contributed by NADH Will not move thru Complex II Cytochrome bcl complex Complex 111 0 11 polypeptide chains in size has Hemes groups 0 2 e are transferred to this complex by Ubiquinone carrier Cytochrome oxidase Complex IV 0 13 polypeptide chains in size 0 2e are transferred to this complex to by Cytochrome C carrier 0 can only transfer one electron at a time cytochrome ala3 inside combines 2 electrons coming off with 2 protons and 12 02 molecule gt H20 0 this detoxifies oxygen 0 Use Cyanide to stop For every 2 electrons that move thru 6 H get pumps across the inner membrane 0 establishes a proton gradient NADH gt6H2e FADH2 gt 4H2e Proton Motive Force Chemiosmotic gradient H are in the inner membrane space that get pumped thru ATP Synthase ATP Synthase 0 F0 region 0 F1 region I synthesizing portion that makes ATP I ATPase 0 For every 2H pumped thru an ATP is made 1 NADH gt 2e gt6H O 3 ATP generated Glycolysis 2 NADH H gt 4 ATP in ETC 2 ATP in SLP PDH 2 NADH H gt 6 ATP in ETC Krebs 6 NADH H gt 18 in ETC 2 FADH2 gt 4 ATP in ETC 2 ATP equivalents Terminal Electron Acceptor Oxygen in Humans but some Prokaryotes use Sulfur Cytochromes change color as the electrons come thru them M Proton Motive Force drives ATP production ATP is in accumulated in the matrix in gets pumped out of the mitochondria by a transport protein with no energy expended Coupled Mitochondria movement of H gt ATP slows down glycolysis and kreb cycle inhibitor Uncoupled Mitochondria movement of H x gt ATP uncoupling protein allows H to move back into the matrix wo producing ATP Swordfish have uncoupling proteins so they can dive in deep water and maintain a high body temperature Brown Fat brown due to the presence of a lot of mitochondra in infant in the back of their neck to maintain proper body temperature heat generation because 0 glycolysis runs at a much higher rate because there is not ATP to inhibit the process have the uncoupling protein 1 UCP1 in mitochondria also in BATS in cold temp increase brown fat in adults Dinitropehnol DNP artificial uncoupler efficient at short circuits use forweight loss Drugs that target mitochondria oligomycin AB stop ATP synthase Cyandie Azide Carbon Monoxide CO stops cytochrome oxidaseComplex 4 Rotenone affects NADH Dehydrogenase Complex 1 0 bottom layer of ponds to push fish up to the surface DNA Deoxyribonuleic Acid blue prints for organisms AGC T Deoxyribose structure The phosphate connects to Carbon 3 on the adjacent DNA molecule The nucleotide attaches to the Carbon 139 Double helix held together by hydrogen bondsweak 1 turn is 104 nucleotides 0 each turn is 34nm between turns Genes DNA that when expressed produces a product Compact Form allow access to genes for expression protect from damage vulnerable when in a linear form 0 Histone Proteins Eukaryotes O O O 0 DNA histone other proteins gt Chromatin H1 H2a H2b H3 H4 2 H2A 2 H28 2 H3 2 H4 gt octameric protein nucleosome H1 histone protein linker I this included with the nucleosome is called a Chromatosome Eukaryotes histone proteins DNA Replication Conservative Replication o conserve parental strand Dispersive Replication 0 end product made up of parent and new strand parent not conserved Semiconservative Replication 0 one parent strand and one new strand Prokaryotic Circular DNA Replication Only one Replication Origin 0 Bubble Forms Eukaryotic Linear DNA Replication More than one replication origin on each chromosome greater number in rapidly dividing cells 300 base pairs in length rich in Adenosine and Thymine several thousandchromosome enzyme target these origins to begin replication 0 ex helicase EnzymesStructures Involved in Eukaryotic DNA Replication Helicase opens helix 1000bpsec Ssbinding proteins hold helix apart 0 RNA primase makes a 3 end for DNA pol to bind to 0 DNA polymerase adds DNA nucleotides to RNA primer w3 end Leading Strand Lagging Strand Okazaki fragments DNA ligase connects together okazaki fragments Topoisomerase I o cuts thru one sugar phosphate backbone 0 allows DNA to pivot around the other strand and release tension Topoisomerase ll 0 cuts through both sugar phosphate backbones allowing for untangling Telomerase 0 appears in aging literature 0 enzyme with a RNA template included 0 adds extra nucleotides to prevent chromosome shortening 0 only in eukaryotic cells 0 In most cells telomerase turns itself off I in cancer cells continue to elongate and maintain ends of their chromosomes Proofreading Enzymes Repair Mechanisms Eukaryotes and Prokaryotes Nucleases 0 endonuclease correct internal errors 0 exonuclease correct errors on the ends 0 DNA Polymerase 0 DNA ligase 0 DNA glycosylase cuts base and leaves sugar backbone RNA sugar ribose o Phosphodiester bond at 3 C just like in DNA 3D Single Stranded Structure I Does NOT permanently store genetic information Central Dogma of Life DNA gt RNA gt Protein New thought DNA gt RNA gt Products because RNA can act as an enzyme DNA Transcriptiongt RNA Translation gt ProductsProteins RNA Transcription will produce RNA from DNA only ONE STRAND is used as the Template RNA chain will be complimentary to template 0 except instead of thymine you will have Uracil AU 0 RNA much shorter than DNA Gene Expression Ef ciency 0 Genes DNA that is transcribed high ef ciency going to get many copies of it going to get a Whole bunch of product from it 0 low ef ciency only get one copy get much less product from it Gene Products 0 RNA 0 proteins enzymes 0 structure 0 other products we don t know about Tvnes of RNA mRNA messenger RNA codes for a protein rRNA ribosomal RNA makes a complex with proteins that is used for protein synthesis tRNA transfer RNA transfers amino acids during protein synthesis snRNA small nuclear RNA important in slicing of premRNA has enzymatic function snoRNA small nucleolar RNA chemically modify RNA miRNA micro RNA can regulate gene expression her favorite 0 certain miRNA in plants that we consume can change our gene expression 9MPPP gt1 SiRNA small interfering can turn off gene expression by degrading mRNA 8 hnRNA heterogeneous nuclear RNA covers all others that we don t need to know Prokarvotic RNA svnthesis Transcription RNA polymerase 0 doesn t have to have 3 end to initiate RNA synthesis 0 Sigma factor binds to RNA polymerase 0 This complex scans gene and clamps down on the gene at the initiation sequence Promotor Sequence conserved Coding strand 0 reading in the 5 to 3 direction 0 Sometimes called sense strand Template strand 0 used to make RNA strand 0 Sometimes called antisense strand 0 Read 3 to 5 0 RNA is transcribed 5 to 3 Promotor not part of the gene 0 Location is important 0 Prokaryotes TATA boxGACA box on coding strand Tells polymerase which strand the gene is on Gene starts at nucleotide 1 and anything before that is upstream 1 start of gene Past start of gene gt downstream 10 nucleotides upstream is where TATA box is found 35 nucleotides upstream is where GACA box is found Consensus sequence I Not exactly the same every time but close enough to recognize as promotor I On test will be told exactly how far from start promoter is 0 Christmas tree 0 Prokaryotes attach multiple polymerases with mRNA tails attached Termination sequence 0 Rho protein independent termination I Inverted repeats hydrogen bond to each other and form hairpin loop which physically pulls mRNA off of template 0 Rho protein dependent termination I rho protein moves along RNA transcript I When transcription pauses rho catches up with polymerase and has same properties as helicase and cleaves RNA molecule off I hairpin loop is still made 0 Operon 0 genes for specific function are transcribed together Polycistronic mRNAgene Prokaryotes 1 RNA polymerase 0 mRNA 0 tRNA 0 rRNA Eukaryotes 3 RNA polymerase 0 RNA polymerase I large rRNA 0 RNA polymerase 2 mRNA 0 RNA polymerase 3 I tRNA I 5srRNA I 7scRNA Eukaryotic gene transcription Initiation O Eukaryote promotor TATA box 2030 nucleotides upstream from start CAAT box 7080 nucleotides upstream Don t need both Enhancer region Elongation O 0 Termination 0 Polymerase 2 termination I Posttranscriptional modification I I Occurs in nucleus I UTR untranslated region one on 3 end and on 5 end I Poly A tail I Capping I Methyl G cap on 5 end I Splicing I Spliceosome Made up of I snRNA 7 different proteins Forms lariat and cleaves interon out and splices together exons Alternate splicing I reassembles mRNA in different ways I Interons I Exons intervening sequence of nucleotides expressed nucleotide sequences I mRNA termination sequence 5 AAUAAA3 signals end of gene I Poly A synthetase I Cuts off anything poly 2 added after termination sequence I Adds a poly A tail after termination sequence I Adds several hundred A 0 Polymerase 1 or 3 termination I termination sequence I row of Ts on coding strand 5 TTTTTTTT 3 I Template strand 3 AAAAAAA539 I rRNA 5 UUUUUUU339 regulatory sequences usually far upstream or downstream from promotor 0 Fold in specific conformation and interact with polymerase 0 Enhancer regions mRNA 0 Transcription gt process gt transport 0 First two happen in nucleus 0 then transported to cytosol of cell Monocistronic mRNA gene 0 One gene gt one mRNA gt one polypeptide Comes from gene 0 Coding sequence Gene DNA gt primary mRNA transcript methyl G cap interons no poly A tail gt Mature mRNA transcript methyl G cap no interons poly A tail Example 0 Given I Gene 100 nt in length I 2 interns each 10 nt in length I Mature mRNA has poly A tail that is 100 nt in length 0 From this information we can determine I Mature mRNA will have I two eX0ns that are 40 nt each total of 80 I 100 poly A tail I methyl G cap 1 nt I Total 181 nt Example Test Question A hypothetical immature mRNA has 0 methyl guanosine cap 0 a 20nt 5 untranslated region included in one of the 3 exons 0 a 40nt 3 untranslated region included in one of the 3 exons 0 3 eX0ns 40 nt each 2 introns 100 nt each 150nt polyA tail How many nucleotides was the gene 0 3 eX0ns X 40 nt 120 nt 0 2 introns X 100 200 nt 0 320 total nt in gene Another question How many nucleotides in in the mature mRNA 0 1 for cap 1 nt 0 3 eX0ns X 40 nt 120 nt 0 150nt from poly A tail 150 nt 0 271 total nt in mRNA Interfering RNA RNAi inhibit gene expression 0 triggered by exogenous double stranded RNA a Virus 0 can come from Transposons endogenous can come from organism itself Hairpin loop structures endogenous Classes of RNAi miRNA microRNA usually comes from Hairpin Loops O 22nt in length 0 noncoding 0 targets 3 UTR of exon in mRNA 0 inhibit eXpresssion or destroys mRNA 0 RISC RNA induced silencing CompleX I protein used to make this process occur 0 Functions of miRNA I Apoptosis I cell growth metabolism I neural development I SILENCES OR DESROYS mRNA siRNA small interfering RNA 0 O O 22nt non coding RNA endogenous or exogenous viral infection by virus have potential to be pathogenic can also come from transposable elements intervening nt sequences or even from some protein coding genes DESTROYs complementary mRNA Functions for Endogenous siRNA I antiviral pathways I silences transposable elements I metabolism piRNA piwiRNA come from transposable elements 0 O O O 2030nt arise from single stranded RNA from transposable elements intergenic regions Functions I Maintains genomic integrity I usually functions in gametes to silence transposons I happens in the nucleus of the gametes I all of the other function at the mRNA level in the cytoplasm all been described in Drosophila know they are used to regulate gene expression where they come from and the mechanism of action for each Protein Synthesis mRNA carries information fro protein synthesis the RNA sequence tRNA carry amino acids to ribosome rRNA rRNA proteins ribosomes decode mRNA gt Translation 0 most abundant RNA in the cell 0 all of these RNAs have different structures 3 Main Types of RNA 1 mRNA only RNA that codes 2 rRNA 0 noncoding and serves as structural backbone of the ribosome 0 synthesized in nucleolus 3 tRNA 0 tRNA and rRNA are most common and are noncoding cterigl ribosomgl genes 0 Each rRNA transcriptional unit consists of 0 16s 0 23s 0 5s 0 The transcription of these genes produces a precursor rRNA prerRNA 0 The prerRNA transcribed is 5 16s23s5s 3 With nonrRNA sequences in between 0 The nonrRNA sequences are called spacers Ribonucleases remove the spacers and release three rRNA separate molecules Ribosomal proteins With the three rRNA make the ribosome two subunits Interons code for L and S subunits Mrvotjc ribosomgl genes 0 Eukaryotic rRNA is coded by rDNA genes 0 The rDNA genes are composed of units containing 0 18s58s28s This unit is repeated in eukaryotic genomes 1001000 times 0 The rDNA repeat units get transshipped by RNA Pol I producing a prerRNA with spacers 0 PrerRNA O 5 18s58s28s 339 0 Ribonucleases process the rRNA unit and remove spacers 0 5s rRNA is located in other location than the repeat unit 0 transcribed independently by RNA Pol III 0 The ribosomal proteins and the four rRNA molecules make the eukaryotic ribosome The Nucleus Nucleolus has 2 specific regions 0 granular region O brillar region I transcription M 0 Know main functional areas 0 Acceptor armstem 0 T armloop 0 D armloop 0 Anticodon armloop O O 3nt 1 amino acid Aminoacyl tRNA Synthetase CHARGING of tRNA 0 Specific enzyme will come and hug tRNA molecule and put amino acid onto it 0 this is called CHARGING OF the tRNA I Start with tRNA AA ATP I First Reaction creates an AAAMP tRNA 2pi I Second Reaction creates AAtRNA AMP I attaches to 3 end Translation mRNA gt Protein mchinerv for Translation Ribosomes tRNA 0 mRNA 20 Common Amino Acids 4A3 64 unique codons 0 redundancy in the code 0 some AA that have more than one codon 0 Degenerative 0 First nt is always the same 0 Second nt is most of the time the same 0 Third nt WOBBLE position varies greatly 3 Codons that do not code for anything Nonsense STOP Codons 0 Code is UNIVERSAL enough that Eukaryotic genes can be expressed in Prokaryotic systems 0 Start Codon 0 AUG methionine gt Eukaryotes 0 AUG formylated methionine gt Prokaryotic Translation Steps focusing on Prokarvotes 1 Initiation 0 What we need to start SOS 30S tRNA IFl IF2 IF3 GTP mRNA I mRNA reads from 5 gt 3 end I IF3 binds to 30S which activates it and locks them both into the Shine Dalagarno sequence on the mRNA I Shine Dalgarno sequence 5 UTR of mRNA I IF2 GTP binds to tRNAwfMET and brings it in to bind to start codon on mRNA IFl then binds to 30S This motion causes the large SOS ribosomal subunit to come in and lock in place I the fMET tRNA binds in the P site only I once everything is combined all initiation factors IF fall off I 0 The Large SOS Ribosomal Subunit I A site I all incoming tRNA come in I P site I growing polypeptide chain I E site I eject empty tRNA 0 In Eukaryotic Initiation I involves Kozak sequence I Methionine not formylated methionine is the first amino acid in eukaryotes I Cap binding proteins and the poly A tail play a role in initiating transcription in eukaryotes 2 Elongation 0 Elongation factors come in I Tu temperature unstable I Ts temperature stable I G elongation 0 First Step Tu bound to GTP goes and gets a tRNA wattached AA and it attaches to A site anticodon of tRNA binds to the codon on the mRNA I this step converts GTP gt GDP 0 Second Step Ts regenerates GTP loaded Tu factor to continue to add tRNA 0 Making of Peptide bond I Peptidyl Transferase enzyme in ribosome that catalyzes peptide bond formation I fMET cut off and joins Gly in A site BUT SIMULTANEOUSLY it SHIFTS TO THE RIGHT USING THE G Elongation Factor as Energy I making the tRNA eject out WALkING down the mRNA I Watch the VIDEO I 3 Termination 0 Stop Codon UAA UGA UAG I no corresponding amino acid they code for a releasing factor I RFl matches up to stop codon I RF3 has energy accompanied with it utilizes energy and breaks ribosomal subunits apart and translation is terminated Polyribosomes multiple ribosomes translating one mRNA 5 339 0 tails of protein 0 Christmas tree structure 0 Can be polycistronic or monocistronic Antibiotics that affect Ribosomes small subunit 30 0 tetracyclins O aminoglycosides streptomycin large subunit SOS O streptogramins O erythromycin Cytosol vs Cytoplasm Cytosol What remains after organelles are removed 0 Cytoplasm organelles cytosol Nucleus 0 Contains Nucleoplasm Plasma Membrane 45 nm thick 0 enclose cell 0 de nes boundaries maintains essential differences between cell types separates organelle environments Fluid Mosaic Model 0 O uid dynamic structure phospholipid bilayer I hydrophilic head I hydrophobic tails I gt Amphipathic molecule Animal Cell is 50 Lipid Red Blood Cell is 40 Lipid and 44 is Protein Mitochondria is 24 Lipid and 78 is Protein 1415 KB I Phosphotidyl cholin outer bilayer I phosphotidyl inostilol inner bilayer I phosphotidyl ethanolamine inner bilayer I sphingomyelin outer bilayer nerve cells 388 KB I phospholipids I proteins I cholesterol 2516 KB Whgt Effects Fluiditv Temperature think about oil in refrigerator 0 Increase T Increase Fluidity 0 Decrease T Decrease Fluidity Length of Phospholipid Fatty Acid think about brush bristles 0 The longer the tails Increase Fluidity 0 Short tails Decrease Fluidity Level of FA tail Saturation 0 think about people standing in a row throwing leg out to side 0 Saturated no DB straightcan pack them in very tight I decrease uidity O Unsaturated DB kinked not able to pack very tightly I increase uidity 0 Cholesterol 0 Normal Temperature 37C decreases uidity stability 0 Extreme Cold maintains uidity Membrane Protein Structure 0 alpha helix 0 hydrophobic AA on the outside 0 hydrophilic AA on the inside 0 Functions I transporters through membrane I receptors alpha only I anchor things inside the cell Beta sheet 0 beta barrels I Hydrophobic AA outside I Hydrophilic AA inside 0 Functions I transporters through membrane I anchorage Glycocalyx O carbohydrate matrix is anchored to integral membrane proteins Small Molecules can pass thru membrane easily C02 0 02 Large Molecules cannot pass through Without help Glucose Amino Acids Anything With a charge Ions 0 H C1Na K Membrane TI E SDOI t 1 Passive Transport 0 no energy expended 0 down concentration gradient High gt Low 0 Simple Diffusion I Constant Linear Relationship I 0 Facilitated Diffusion I Channel Protein I Gated 1 Ligand gated I closed until its specific ligand comes in and bindsnot What goes thru opens gate I 2 Voltage gated I seen in neural cells action potential I momentary voltage shift gt gates open I K shifts OUT major intracellular ion I 3 Stress gated I can be opened by sound waves ears I sensory receptor I I Ungated open all of the time but specific I Carrier Protein I advantage specific I disadvantage levels out due to saturation of the binding sites I 2 Active Transport 0 requires energy ATP 0 moves up concentration gradient Low gt High 0 Active Transport w Transporter Protein I Uniport I ATP used I lmolecule 1 direction I I Symport I ATP used I 2 molecules 1 direction I I Antiport I ATP used I 2 molecules opposite directions I Example NaK pump critical for cell survival I Oubain dart frogs block NaK pumps Endocytosis O uptake large molecules 0 uptake specific molecules 3 Phagocytosis O uptake of large nonspecific particles 0 example Macrophages and foreign body I Oligosaccharides on surface of cells help macrophages recognize foreign body I Apoptosis programmed cell death I changes is phospholipids of cells undergoing apoptosis signal macrophagesneutrophils to come and engulf them 4 Pinocytosis cellular drinkingquot 0 ingest small volumes of uid or small molecules I 5 Receptor Mediated Endocytosis 0 Example LDL cholesterol 0 High LDL problem with receptors 0 Cytoskeleton Functions 0 Cell framework 0 Cell motility 0 Cell contraction Cvtosllteleton Macromolecules 1 Microtubules largest alpha tubulin beta tubulin largest cytoskeletal molecules 25m in diameter present in all eukaryotic cells Heterodimer alpha beta tubulin Uniform size and shape Protofilament and end 2 Typically attached to MTOC at ONE end I Microtubule Organizing Center I Stability established by GTP cap on Heterodimer I stable growing microtubule I Microtubule Function I Moving vesicles in cells I cell divisiondifferentiation I cell shape I cell motility Drugs I Taxol anticancer drug stabilizes microtubules so they don t break down in cell division I Colchicine binds tubulin and blocks polymerization I inhibits mitosis I allows for preparation of slides to show condensed chromosomes Microtubule Associated Proteins MAPS I Tau Hyperphosphorylation results in protein tangles associated with Alzheimer39s Disease I Motor proteins walk along microtubules CiliaFlagella in Eukarvoteg l 92 arrangement I 9doublets I central pair I I l Motor Protein in 92 arrangement of MT and Centrioles l Dynein Protein toward nucleus sigmoidal movement like a snake walking toward the end in cilia and agella gives you the functionality WILL HAVE TO DRAW ON THE EXAM Spoke protein I Kineisin Protein towards membrane I moving cellular components I walks toward the end away from MTOC I NOT in cilia or agella I Primary Cilia I not used for motility I receive signals from the environment Microfilaments Actin monomers can occur in cross linked bundles I GActin globular I has ATP attached I FActing fibrous I 67nm in diameter Microfilament Function I muscle actinmyosin I Motility I Drugs I Phalladin from amantida mushroom inhibits actin depolymerization 3 Intermediate Filaments largest heterogenous family of keratin protein fibers 5 Classes I Type I and II acidic and basic keratin filaments I EX Hair and Nails I Type III I sarcomeres of muscle I glial cells astrocytes I Type IV I neurofilaments I along axons l Type V I lamins right under phospholipid bilayer Endonlgsmic Reticulum makes up 12 of all membrane in the cell 0 highly branched structure 0 continuous With the outer nuclear membrane 2nd phospholipid bilayer 1 Rough ER RER 0 Ribosomes attached 0 Protein synthesis Membrane proteins I EX Integral proteins Proteins for other organelles I EX Mitochondrial proteins Proteins for ER Proteins released from cell When signal peptide sequence during translation in the cytosol is recognized by signal recognition protein translation stops and the ribosome mRNA and polypeptide are brought to RER It attaches by docking to protein translocation protein and translation begins Once it reaches a stop codon the polypeptide is cleared at the signal sequence and it is translocated thru into the lumen of the RER I Happens for I ER protien I Secreted protein If you want to integrate this protein into the membrane I Beginning of mechanism is the same but as proteins Polypeptide moves into lumen certain sequences are hydrophobic Which causes hinge to open in translocation protein and the strand of polypeptides are integrated into membrane I gt single pass membrane protein I this happens 7 times for a normal 7 membrane pass I I I Proteins Will bud off from ER and migrate up to the cell membrane and fuse With membrane Then opens up bud and is now integrated into the plasma membrane Whatever is inside vesicle that leaves the RER will be on the outside of the cell membrane I Proteins are started on NH2 end and grow on COOH end Transmembrane glycosylation I Sugar addition to proteins made in RER I There is a particular amino acid sequence that signals time to attach sugar I asn asparagine AA thr or ser I This is the sequence that signals sugar addition I Dolichol I Modified lipid I Has ability to build sugars off of it I Sugars from this get attached to asn I I Membrane proteins can be attached through sugars to lipids 2 Smooth Endoplasmic reticulum SER I Phospholipid synthesis I Phosphotidyl choline I Major lipid in outer bilayer I gt 50 I All synthesis happens in cytosolic side of membrane I Enzymes will ip phospholipids across the bilayer I Scramblase I Randomly ips any phospholipids I Flippase I Flips phospholipids With free amino groups phosphotidylserine ethanolamine etc I 2 fatty acids glycerol polar head I Steroid hormone synthesis I Detoxification I Many drugs that are water insoluble are detoxified here I Cytochrome p450 I Sequesters calcium I Sarcoplasmic reticulum is modification of SER I Organelle membranesproteins Golgi Lysosomes endosomes All of these come from ER I Exception is mitochondria I Phospholipids are transferred from SER to mitochondria by a phospholipid transfer protein 0 Smooth ER SER O tubular appearance 0 Phospholipid synthesis 0 Steroid Hormone synthesis 0 Drug Detoxification enzyme Cytochrome p450 0 sequestering of CaZ Golgi Structure location near nucleus numerous membranebound discs gt Cisternae 6 Cisternae Golgi Stack More than one Golgi in a cell produces a lot of products that get transported out of the cell Specific Orientation Cis face closest to the nucleus bumpy appearance due to vesicles coming in an binding gt Fenestrate Convex Trans face furthest from the nucleus Transport forms vesicles leaving the cell Concave Vesicle Release Constitutive Release gt Continuous Regulated gt Requires a Signal ex insulin Golgi Function Glycosylation modifying sugars ln RER Glycosylation nlinked attached to NH2 ex asparagine ASN ln Golgi Glycosylation olinked attached to Hydroxyl group OH ex serine threonine Sugar Transport into Golgi Galactose Fructose Sialic Acid Mannose nacetyl glucosamine Most common sugar in nature Pro Processing activating proteins Membrane Recycling Endosome PIC Lysosome 50 hydrolytic enzymes Add Notes from blake primary lysosome secondary lysosome primary whatever it binds Func ons intracellular digestion Digestive vacuole Autophagy Autophagic vacuole digestive vacuole autophagic vacuole residual body extracellular digestion Specialized Lysosomes sperm acrosome secrete hydrolytic enzymes for egg penetration egg breaks egg yolk protein so it can be consumed by embryo Peroxisomes Break down Fatty Acids Beta oxidation and Amino Acids Contain oxidase enzymes and catalase enzymes humans lack uric acid oxidase reason we get Gout Major oxygen consumers Oxidase Reaction RH2 02 gt R H202 RH2 phenol formic acid toxic compounds Catalase Reaction Use Catalase to break down H202 H202 R H2 gt 2H2O R39 R H2 another toxic compound produces 02 also only in prokaryotes Mitochondria Contain DNA Contain Ribosomes more viscous cytoplasm Storage Site for CALCIUMl Matrix Cristae lnner Membrane high surface area due to cristae less permeable Proton gradient ETS is here transport proteins inner membrane resembles prokaryotic membrane ex Cardiolipin modified phospholipid found in bacteria and mitochondria Higher protein content Outer Membrane 40 lipid more permeable than the inner membrane similar to eukaryotic membrane contain PORIN proteins Beta Barrels allow movement across membrane 2 3nm 1213000 mitochondria in liver cell Functions of Nucleus Houses DNA Regulates transcription Every cell in body has full amount of DNA for entire organism What makes cells unique is differential gene expression Once a cell has gone down this path it cannot go back All cells have common structures but have different levels of them depending on what kind of cells they are Each cell type has unique set of proteins that are specific to that cell type Average cell 2000 abundant proteins Only certain genes need to be expressed at certain times Prokaryotic cells Use operons to control gene expression Cluster of genes controlled by a single regulatory signal Eukaryotic cells Use promotors and transcription factors to regulate gene expression 7 o of genes expressed in average cell Positive regulation is more efficient Everything is off until it is turned on This way something is only made when it is needed Transcription factors Regulatory region of genes bound by transcription factors Zinc finger Helixloophelix Homodomains Allow polymerases to bond better Also affect how much transcription happens Master regulatory Gene MRG its protein product can go and turn on other genes which in turn turn on more genes cascade effect Example of this is when an embryo is developing and cells are differentiating Critical genes for determining what you will get in the end Only need a small number of them to have a huge amount of individual cell types Turned on by transcription factors Transcription factor III A TFlllA Controls 5srRNA transcription Each cell has both Somatic type 5srRNA genes Somatic cells express this type 350 copies of gene Oocyte type 5srRNA genes Oocytes express this type Aournd 19000 copies of gene How tightly the TF binds to internal control region determines whether oocyte or somatic type of 5srRNA gets expressed Somatic cell high affinity for TFlllA lower levels of TFlllA Most of the TFlllA will get caught by the somatic genes Oocyte cell Increased amount of TFlllA There will still be some somatic type 5srRNA expressed since it still has a higher affinity Once levels of 5srRNA become sufficient they bind to the TFlllA and create 7srRNA inactivated TFlllA Within 5srRNA gene there is an internal control region In between nucleotides 53 and 83 This is where TFlllA binds Forms of gene expressionProtein Expression Prokaryotes Control through Transcription Only Eukaryotes Different levels of protein expression DNA modification Epigenetics Methylation Acetylation these both space out histones allowing DNA polymerase to bind increasing transcription Histone modifications Chromosome inactivation Packing in DNA so tightly that it cannot be reached Transcriptional Master regulatory genes Transcription factors Enhancers Post transcriptional GCap Poly A tail Remove introns Translation Number of ribosomes More ribosomes more proteins ex polyribosome Translation factors Stability of mRNA Post translational Proprocessing Phosphorylation Methylation Compartmentalization lnterphase nucleus Not actively dividing Has outer and inner phospholipid bilayer Space in between 2040 nm Can add membrane from ER Each membrane is specially designed to interact with either the cytoplasm of cell or the nucleoplasm DNA inside nucleus is in form of chromatin DNA proteins chromatin Heterochromatin condensed DNA Euchromatin dispersed DNA Ribonucleic proteins RNP Process newly formed mRNA Nucleioribosome synthesis Nucleolus Transcription of rRNA occurs fibrillar Assembling of rRNA and proteins granular Proteins associated with nuclear membrane Nuclear pores Nuclear pore passes through both bilayers 100 nm across total diameter 9 nm opening 30004000 pores on surface of nucleus All big traffic in and out of nucleus moves through these pores Not sure how nuclear basket allows things to pass through ribosomes mRNA will move through these Polymerases and proteins ie RNA proteins need to come into nucleus Know structure Nuclear lamina Mechanical support Foundation for phospholipids to give nucleus shape Also helps hold pores in place anchoring Keeps DNA from flowing out of pores Can be tethered to lamina 5080 nm thick Lamins A B and C make up lamina Most abundant DNA binding proteins in cell Theories to explain second membrane on nucleus Syntrophic model Hypothesis that nucleus was created through symbiotic relationship bn bacteria and archaea Could explain double membrane Viral Eukaryogenesis hypothesis Idea that eukaryotic cells arose from viral attack of prokaryotic cell Viral DNA and membrane would have become nucleus Viral DNA and Eukaryotic DNA are both linear Exomembrane hypothesis Extra membrane arose from mutation Membrane Function Absorption Secretion Fluid Transport Mechanical Attachment Cell Communication Cell Adhesion Molecules CAMS Bind to each other as well as intracellular proteins Form different types of cellular junctions 4 Major Families Cadherins lmmunoglobins superfamily of CAMs lntegrins Selectins Cell Junctions You can have multiple connections on one cell Anchoring junctions 3 Main Types Adherens junction actin and cadherin protiens adhesion belt actin filaments factin help anchor cadherin proteins seen in epithelial cells cell attachment thick of like velcro Desmosomes a single point of attachment have cadherin proteins that interdigitate btw 2 cells that are mediated via CALCIUM attached to protein plaques which is anchored by intermediate filaments seen in smooth muscle strong connections Hemidesmosomes half a desmosome bottom of cells anchor cells to basal lamina protein matrix associated with protein plaque instead of cadherin protein there are INTEGRINS Keratin Filaments Occluding junctions keep things from moving backforth Tight Junctions Vertebrates Alimentary canalIntestines makes seal btw cells on apical end quilting pattern Proteins involved Occludin and Claudin Septate Junctions Invertebrates Channel junctions help to move molecules backforth Gap Junctions animals Heart Muscles some neurons complex tunnel thru membrane protein involved Connexin 6 make up channel called Connexon Plasmodesmata plants Signal Relay junction not in direct contact but exchange signals for communication Chemical synapses immunological synapses transmembrane ligand receptor cell cell signaling contacts GET A CHANCE TO EARN BONUS POINTS ON NEXT EXAM Nucleolus DNA Fibrillar Euchromatin in Nucleolous loosely packed and being transcription of tandem repeats of rRNA 185828 Granular rRNA assembly into ribosomal subunits RNA Protein Also in nucleolus there is Heterochromatin Facultative Barr Body Constituitive Never expressed always condensed ex Structure
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