intro to cell & molecular biology how do we study cell biology? –reductionist view cells as...
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Intro to Cell & Molecular Biology• How do we study cell biology?
– Reductionist view• Cells as tiny complex machines• Sum of parts = whole
• Your goal:– be able to explain the roles
various molecular parts play in cell biological processes
– With the same clarity as with macroscopic items (bicycles, stoves, trains, etc…)
Intro to Cell & Molecular Biology• How do we study cell biology?
– Parsimony• the simplest explanation for all
relevant data is preferred over more complex explanations
• The most parsimonious answer is not necessarily perfectly correct
• More data could make us revise it
Chemical basis: Bonding• Covalent Bonds: sharing of e-
– One pair shared = single bond C C– Two pairs = double bond C C– Three pairs = triple bond C C
• Electronegativity (EN) is the ability of an atom to attract electrons to itself– C = 2.5 N = 3.0 O = 3.5 H = 2.1 S = 2.6– Sharing is unequal between different atoms in a molecule
• Polar molecules have significant EN differences– H2O, CH3COOH
• Nonpolar molecules have little EN differences– CH3(CH2)nCH3
• Amphipathic molecules have different EN characteristics at different positions
– CH3(CH2)nCOOH
Chemical basis: Bonding• Noncovalent Bonds: attractive forces between atoms of
opposite charge– Ionic: fully charged Na+ Cl-
• Strength dependent on environment (salt crystal vs aqueous)– Hydrogen: partial charge (polar molecules)
Noncovalent bonding• Noncovalent Bonds: continued…
– Van der Waals: transient dipole interactions
– Hydrophobic: water fearing– Hydrophilic: water loving
Robot Lizards ExploitVan der Waals contacts
• Based on the Gecko• Adhesion depends on close
contact between surfaces “StickyBot”
• Video link
H2O• Can form 4 hydrogen bonds
– High energy barrier to liquid --> gas phase
transition
• Highly polarized– Asymmetric structure - both H atoms on one side– Can dissolve many compounds
H2O• Can dissolve many compounds
– Acids: can release H+– Bases: can accept H+
pH = - log [H+]• Pure H2O pH = 7 , [H+] = [OH-] = 10-7 M
• Why are reactions so pH sensitive?– Amino acid functional groups can change state based on pH
Carbon• Central to the chemistry of life.
– Can form four covalent bonds, with itself or other atoms.
– Carbon-containing molecules produced by living organisms are called biochemicals.
Chirality and Stereoisomerism• Chirality and Stereoisomerism:
– Asymmetric carbons bond to four different groups.
– Two mirror-image configurations:• Enantiomers, (aka)• Stereoisomers
– Can be either D- or L-isomers
– Natural amino acids = almost all L-isomers
– Natural carbohydrates = almost all D-isomers
Classes of molecules• Miscellaneous co-factors
– Vitamins, ATP, NADPH, etc
• Metabolic intermediates– Glycolysis, TCA cycle, etc
• Monomers– Amino acids– rNTPs = A, G, C, U– dNTPs = A, G, C, T– Sugars
• Macromolecules
Classes of molecules• Macromolecules
– Lipids• Fats = glycerol esterified with 3 fatty acids
– Saturated, unsaturated, cis, trans• Phospholipids = glycerol + 2 fatty acids + 1 phosphate• Steroids = cholesterol and derivatives
Classes of molecules• Macromolecules
– Lipids• Fats = glycerol esterified with 3 fatty acids
– Saturated, unsaturated, cis, trans• Phospholipids = glycerol + 2 fatty acids + 1 phosphate• Steroids = cholesterol and derivatives
Classes of molecules• Macromolecules
– Lipids• Fats = glycerol esterified with 3 fatty acids
– Saturated, unsaturated, cis, trans• Phospholipids = glycerol + 2 fatty acids + 1 phosphate• Steroids = cholesterol and derivatives
Classes of molecules• Macromolecules
– Lipids• Fats = glycerol esterified with 3 fatty acids
– Saturated, unsaturated, cis, trans• Phospholipids = glycerol + 2 fatty acids + 1 phosphate• Steroids = cholesterol and derivatives
Monomers and polymers
Classes of molecules• Macromolecules
– Carbohydrates• ( CH2O )n• At n ≥ 5 self-reaction to form rings
– C5 = ribose monomer– C6 = glucose monomer
Classes of molecules• Macromolecules
– “Nutritional” sugars:» Glycogen = branched alpha 1-4 linkage, dense granules
in cell cytoplasm in animals» Starch = helical and branched alpha 1-4 linkage, within
membrane bound plastids in plants
plastid
Classes of molecules• Macromolecules
– “Structural” sugars:» Cellulose = long and unbranched, beta 1-4 linkage,
resist tensile (pulling) forces, plants» Chitin = unbranched, N-acetylglucosamine,
invertebrates» Glycosaminoglycans = components of extracellular
matrix for cartilage and bone, repeating (A-B)n structure
Classes of molecules• Macromolecules
– Nucleic Acids• Nucleotide monomers (rNTPs, dNTPs)• Storage and transmission of genetic information
– Phosphate + 5C ribose sugar + nitrogenous base
RNA DNA
H
• DNA is usually double stranded• RNA is usually single stranded
– RNA may fold back on itself to form complex 3D structures, as in ribosomes.
– RNA may have catalytic activity; such RNA enzymes are called ribozymes.
– Adenosine triphosphate (ATP) is a nucleotide that plays a key role in cellular metabolism
– Guanosine triphosphate (GTP) serves as a switch to turn on some proteins.
Classes of molecules
DNA is a useful reference-frame for sizes
Classes of molecules• Macromolecules
– Proteins• Amino acid monomers• Peptide bond formation• N-terminus versus C-terminus• Backbone is common, side chains (R) differ
Classes of molecules• Macromolecules
– Proteins• Backbone is common, side chains differ
– 4 categories of amino acid side chains» Polar charged D, E, K, R, H» Polar uncharged» Nonpolar» Unique
Classes of molecules• Macromolecules
– Proteins• Backbone is common, side chains differ
– 4 categories of amino acid side chains» Polar charged D, E, K, R, H» Polar uncharged S, T, Q, N, Y» Nonpolar» Unique
Post-translational modifications: Phosphorylation of –OH groups
Classes of molecules• Macromolecules
– Proteins• Backbone is common, side chains differ
– 4 categories of amino acid side chains» Polar charged D, E, K, R, H» Polar uncharged S, T, Q, N, Y» Nonpolar A, V, L, I, M, F, W» Unique
Classes of molecules• Macromolecules
– Proteins• Backbone is common, side chains differ
– 4 categories of amino acid side chains» Polar charged D, E, K, R, H» Polar uncharged S, T, Q, N, Y» Nonpolar A, V, L, I, M, F, W» Unique G, C, P
Hydrophobic and hydrophilic amino acid residues in the protein cytochrome c
Levels of protein structure• Primary
– Sequence of the polypeptide chain
H3N-MQWERTYIHAHAPKLCVN-COOHH3N-Met Gln Trp Glu Arg Thr Tyr Ile…
H3N-Methionine Glutamine Tryptophan…
Levels of protein structure• Secondary
– Alpha-helix (collagen)– Beta-sheet (spider silk)– Side-chain dependence to which form is adopted but stabilization
comes from backbone - backbone hydrogen bonding interactions
Levels of protein structure• Tertiary
– Side-chain dependent and mediated packing of the secondary elements– Fibrous proteins = elongated, often structural roles– Globular = compact, often enzymes
Protein domains can be modular
• Protein Domains– Domains occur when
proteins are composed of two or more distinct regions.
– Each domain is a functional region
Protein structures can be dynamic
• Dynamic Changes within Proteins– May occur with
protein activity.
– Conformational changes are non-random movements triggered by various events (e.g. binding, chemical mods…)
Levels of protein structure• Quaternary
– Interactions between 2 or more distinct polypeptide chains
• Protein-Protein Interactions– Results from large-
scale studies can be presented in the form of a network.
– A list of potential interactions can elucidate unknown processes.
Disease• Sickle-Cell Anemia (SCA)• Painful• Life-threatening
periods of crisis• e.g. vaso-occlusive
crisis• block blood flow
in capillaries
Disease• Sickle-Cell Anemia (SCA)
• Hemoglobin is composed of four polypeptide chains• Two alpha-globin subunits + two beta-globin
subunits• SCA is caused by a single amino acid substitution
in beta-globin• E6V
Protein structure and folding
• Anfinsen RNase A experiment– Denature (unfold) protein in urea• Observed loss of activity
– Dialyze the urea away• Observed refolding• Regain of activity
Demonstrated structural information is inherent to protein sequence• Follow a folding pathway• Fold to the lowest energy state
Two alternate pathways for protein folding
Chaperones prevent mis-folding• Molecular Chaperones
– HSP70 during translation of nascent peptide• Binds exposed hydrophobic regions• Hydrolyzes ATP in a bind-release cycle
Hartl, et al (2011)
Chaperones prevent mis-folding• Molecular Chaperones
– HSP70 during translation of nascent peptide• Binds exposed hydrophobic regions• Hydrolyzes ATP in a bind-release cycle
– Chaperonins assist post-translation
Protein folding and Disease• CJD (Mad Cow) & Alzheimers Disease
– PrPC --> PrPSc --> plaque– APP --> Ab42 --> plaque