Prelim 1 Slides F24 PDF

Prelim 1: Next Monday, Sep 23th All exams are taken on CANVAS: CANVAS/EXAM/Exam Instruction Check your seat assignment (by Friday Sep 20th): CANVAS/EXAM – either in Call Auditorium or Klarman G70. Full instructions on Canvas/Exams/Exam Instructions....

Prelim 1: Next Monday, Sep 23th All exams are taken on CANVAS: CANVAS/EXAM/Exam Instruction Check your seat assignment (by Friday Sep 20th): CANVAS/EXAM – either in Call Auditorium or Klarman G70. Full instructions on Canvas/Exams/Exam Instructions. It is your responsibility to read and follow the instructors. Not following the instructions can be considered cheating and may result in 0 points in the Prelim and further penalties. So please don’t!!! Material: lectures 2-7 and sections 2-4 Note: we grade based on material taught in class (lectures and sections) Practice Prelim 1 is available on: CANVAS/EXAMS TAs review sessions! Saturday/Sunday 1-4 PM Biotech Racker Room G01 (Instructions in CANVAS/EXAMS) Office Hours with Martin Graef: Mon 5-6 pm Biotech 201 Wed 1:30 – 2:30 pm Biotech 201  Proteins in Action Learning Objectives: Understand how the specificity in binding interactions is achieved Understand how enzymes catalyze favorable and unfavorable reactions. Understand mechanisms by which proteins are regulated, including allostery and phosphorylation Understand how “small GTPases” transmit signals as molecular switches Different ways to depict a protein structu Different ways to view a protein structure TODAY’S TOPICS: 1.Proteins as Enzymes 2.Regulation of Protein Activity The “Central Dogma” of Molecular Biology nucleus DNA sequence mRNA sequence cytosol Protein sequence “compartmentalization”  Protein structure can be described in 4 levels of organization Primary Secondary Tertiary Quaternary structure amino structure a- structure 3D structure 3D acid sequence helices & b- structure of a structure of sheets polypeptide multiple amino acids chain polypeptide chains N- to C-terminus 3D structure allows proteins to execute particular functions Glu35 Asp52 P00698 · LYSC_CHICK ective binding of a protein to a ligand depends a set of weak noncovalent interactions and interactions (H-bonds, van der Waals, electrosta and hydrophobic interactions) A substance (ion, small organic molecule, or macromolecule) that is bound by a Figure 4-27 Essential Cell Biology  Binding sites in proteins interact with very specific ligands cyclic AMP (ligand) Folding into the 3D protein structure ideally positions critical sidechains in a way that allows for very specific non-covalent interactions (here hydrogen bonds and Electrostatic interactions) with the ligand (and not oth Figure 4-28 Essential Cell Biology  Enzymes catalyze (speed-up) chemical reactions while remaining unchanged themselves chemical active reaction site “substrate (S)” “product” or “reactant” “ES” “PS” Figure 3-15 Essential Cell Biology  Enzymes catalyze reactions by lowering the activation energy barrier “transition state” “transition state” substrat substrat e (S) e (S) product product (P) (P) Figure 3-12 Essential Cell Biology  Enzymes catalyze reactions by lowering the activation energy barrier Activation energy barrier Figure 3-14 Essential Cell Biology  Carbohydrates (sugars and saccharides) monomer macromolecule monosaccharide polysaccharide (dozens) monomer glucose “Carbohydrate” = carbon + water Like DNA and RNA, sugars can form long Polymers by condensation reactions Chemical formula: (CH20)n (n = 3, 4, 5 or 6)  Carbohydrates (sugars and saccharides) monomer macromolecule monosaccharide polysaccharide (dozens) 1 Monosaccharide eg. glucose, fructose, ribose, deoxyribose, etc 1 2 Disaccharide eg. sucrose (“sugar”), lactose, maltose, etc ~3-10 copies Oligosaccharide >10 copies Polysaccharide eg. glycogen, starch, cellulose, chitin  Polysaccharides Glycogen (up to ~50,000 glucose monomers) branch points Functions of saccharides 1. Energy storage (glycogen), structural support (cellulose* in plants), extracellular matrix in animals, and cell walls in bacteria and fungi 2. Protein and lipid modification (“oligosaccharides”) * Cellulose is the most abundant macromolecule on earth because about 1/3 of plant mass is cellulose! Panel 2-3 Essential Cell Biology  Hydrolysis – the opposite reaction to condensation polysaccharide H2O + “hydrolysis” If hydrolysis is energetically favorable, why are macromolecules (DNA!) stable in ce Shouldn’t they all spontaneously fall apar substr ate produ ct  Lysozyme can cleave polysaccharides polysaccharide H2O + “hydrolysis” Activation energy keeps macromolecules sta and prevents spontaneous hydrolysis! Imaging what would happen to your DNA substr if the activation energy was very low! ate produ ct Figure 4-40 ECB6  Lysozyme can cleave polysaccharides polysaccharide H2O + “hydrolysis” Activation energy keeps macromolecules sta and prevents spontaneous hydrolysis! Imaging what would happen to your DNA substr if the activation energy was very low! ate Using enzymes to lower the activation ener when produ needed allows cells to regulate proces ct Figure 4-40 ECB6  Lysozyme can cleave polysaccharides (Lysozyme helps to hydrolyze the polysaccharides in bacterial cell walls) polysaccharide H2O + “hydrolysis” lysozyme polysaccharide Substrate Enzyme- Enzyme- Product + Substrate Product + Enzyme complex complex Enzyme Figure 4-40 ECB6 3D structure allows proteins to execute particular functions Glu35 Asp52 P00698 · LYSC_CHICK Here is the actual structure of lysozyme from chicken  How does lysozyme lower the activation energy? H2O “hydrolysis” S+E ES EP E+P  Here is the actual structure of lysozyme from chicken Glu35 Asp52 P00698 · LYSC_CHICK overall structure has evolved to place Glu35 and Asp52 in the right posi Substrates do not fit into enzymes Enzymes bend and like a key into a lock break the key!  Enzymes speed up reaction rates, o not change whether a reaction is favorable o energetically favorable reactionenergetically unfavorable reaction substrat product e (S) (P) product substrat (P) e (S) Enzymes cannot change the relative energies of substrates and products! cells synthesize all these polymeric macromo if a condensation reaction is unfavorable? energetically unfavorable reaction monomer polymer DNA, RNA, protein, polysaccharides monomer  Reaction “coupling” can drive unfavorable reactions reaction 1: reaction 2 energy +23 kJ/mole reaction 1 reaction is unfavorable and will not occur reaction 2: -energy energy -30.5 kJ/mole +energy reaction is highly favorable coupled reactions catalyzed within an enzyme energy -7.5 kJ/mole coupled reaction is favorable the most widely used chemical energy source ATP hydrolysis releases lots of energy “hydrolysis” = (a molecule of H2O breaks a bond) (ATP = Adenosine tri-phospha +energy -energy gy released by ATP hydrolysis is coupled to hundreds of reaction Figure 3-31 Essential Cell Biology Where did the energy come from during RNA synthesis  Nucleotides in RNA form polymers by phosphodiester linkage monomer monomer dimer condensation H 2O 5’ 5’ 3’ condensation 3’ OH OH 5’ H 2O + 5’ O- 3’ OH 3’ OH Panel 2-6 Essential Cell Biology here did the energy come from during protein synthes acyl-tRNA synthetases attach amino acids to Hydrolysis of ATP to AMP + O- TODAY’S TOPICS: 1.Proteins as Enzymes 2.Regulation of Protein Activity fferent ways of regulating protein activity in ce 1. Control of protein amount: Rate of mRNA transcription (gene expression) Rate of mRNA degradation Rate of mRNA translation into protein Rate of protein degradation 2. Control of protein activity: Cellular localization (e.g. targeting to nucleus) Inhibition or activation by ligands (e.g. feedback inhibition) Inhibition or activation by another protein Protein modification (e.g. phosphorylation)  Feedback inhibition in metabolic pathways A F Product F inhibits enzyme 1 How does this work at the molecular level? Inhibition occurs through “allostery” => “conformational change” allosteric ligand Figure 4-44 ECB  Phosphorylation regulates protein activity Substrate’ protein OR osphorylation is a very common way of regulating protein functi => Changes how the protein interacts with other molecules! Figure 4-38a Essential Cell Biology Regulation by phosphorylation is very common N C kinase domain 518 kinases in humans (~2% of proteome) Each kinase has one or many substrate (target) proteins Kinases often phosphorylate other kinases in signaling pathways ~200 phosphatases in humans >51,000 phosphorylation sites in the human proteome  Small GTP-binding proteins are molecular switches (aka “GTPases” or ”monomeric GTPases”) GUANINE GDP “GEF” Guanine GTP hydrolysis nucleotide “GAP” Exchange GTPase Activating Factor Protein GTP binding GUANINE GTP bind “effector” proteins to activate a downstream pathway There are over 100 ‘small GTPase’ proteins in the Human Genome Growth Control Regulation of the Membrane Cytoskeleton Traffic Membran Traffic thru e Nuclear Traffic Pores Each ‘small GTPase’ regulates a downstream pathway Ras, an oncogene, was the first ‘small GTPase’ to be characterized… The Ras GTPase regulates cell growth ‘Growth GUANINE Ras factor’ GDP signal GTP hydrolysis Ras The Ras G12V GTP binding mutation leads to * cancer because the Ras protein cannot hydrolyze GUANINE GTP and Ras is therefore always active GTP Cell growth signal