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BIBC 100 Midterm 2 Review Session. (Midterm Tomorrow In Class). DNA. B-DNA Right handed antiparallel double helix Purines AG, Pyrimidines CT (U in RNA) G = C, A=T hydrogen bonds Negatively charged 2’ deoxyribose backbone A-DNA Dehydrated, compressed
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BIBC 100Midterm 2 Review Session (Midterm Tomorrow In Class)
DNA • B-DNA • Right handed antiparallel double helix • Purines AG, Pyrimidines CT (U in RNA) • G=C, A=T hydrogen bonds • Negatively charged 2’ deoxyribose backbone • A-DNA • Dehydrated, compressed • Deep, narrow major groove not easily accessible • Z-DNA • Left-handed • Alternating N/M sequences • Tm • Melting temperature at which 50% are SS and 50% are DS • Higher G=C content leads to higher Tm
RNA • A lot more common than DNA • Used in transcription, translation • mRNA (messenger) • mRNA transcript from DNA template • tRNA (transfer) • Attached to an amino acid at CCA 3’ end by amino acyl synthetase • Elongates protein chain in translation at the ribosomes • rRNA (ribosomal) • Makes up structure of ribosomal subunits
RNA • Can form secondary structures/motifs • Stem-loop • Hairpin • Pseudoknots • DNA/RNA and RNA/RNA double helices have A-DNA conformation • Can carry out enzymatic and regulatory activities • mRNA has 5’ 7-methylguanosine cap, but translation starts at first Met (AUG) codon • tRNA carries an anti-codon that wobble-base-pair with the codon (20 aa, 4^3 possible codons)
DNA-Binding Proteins • Bind to the major and minor grooves of B-DNA • Major groove 12A wide 8A deep, binds alpha helices (specific surface pattern for each AT, TA, GC, and CG base pair) • Minor groove 5A wide 8A deep, binds beta sheets (less specificity due to TA/AT and GC/CG base pairs having same exposed surfaces) • Exposed groups • H-bond donor, H-bond acceptor, Hydrogen (VdW), Methyl (VdW) • Motifs • Helix-turn-helix • Zinc finger (Zinc prosthetic group) • Leucine Zipper (heptad coiled-coil hydrophobic stabilization above DNA-Binding domain)
DNA-Binding Proteins • Examples • EcoR1 restriction enzyme (GAATTC palindromic) • Lac repressor in e.coli (actively binds to DNA to prevent txn of lactose metabolism proteins when no lactose is present) • TATA Box Binding Protein (binds TATA box, unwinds DNA, recruits RNA Polymerase to transcribe mRNA from DNA)
DNA Organization • Interphase Chromatin • Unraveled DNA and associated proteins (histones, scaffolds) • Double stranded, but no duplicates • Two types: euchromatin (uncondensed, actively transcribing) and heterochromatin (condensed, repressed) • Mitotic Chromosomes • Condensed heterochromatin, compact, transportable, four arms • Each chromosome has 2x sister (identical) chromatids attached at centromere • Humans have 22 autosomes and 1 sex chromosome (X or Y). Each has its homologous (related in genes, but not identical in content) chromosome, making for 46 total chromosomes in the human diploid (2n) genome
Histones • Interphase chromosomes are organized as 10nm beads-on-a-string conformation • DNA are spaced out with histones attached at regular intervals • 2 winds of DNA around a histone octamer core, with H1 histone linker on the outside (H1 linking creates solenoid structure) • Structure • Core histones 2x (H2A, H2B, H3, H4) • Alpha helical protein • C-terminal covalent modifications • Phosphorylation, monoubiquitination (signaling) • Acetylation (expression) • Methylation (repression)
Sugars • Monomers • alpha-D-glucose (dextrose) • Aldoses, ketoses • Cyclization into pyranose • Dimers • Sucrose (glucose a(1->2) fructose) • Lactose (galactose b(1->4) glucose) • Maltose (glucose a(1->4) glucose) • Anomeric carbon • Attached to anomeric hydroxyl • Either alpha (down) or beta (up) in Haworth • Alpha is cis to D (4’) hydroxyl (same right side in Fischer) • Beta is trans to D (4’) hydroxyl (opposite sides) • Animals cannot break beta glycosidic bonds (but bacteria can)
Sugars • Polymers • Homo/hetero polysaccharides • Glycosidic bond • Alpha – helical structure, compact, energy storage • Animals, bacteria use glycogen glu (1->4) glu – branched every 8-12 res • Plants use starch glu a(1->4) glu • Amylose – unbranched • Amylopectin – branched every 24-30 res • Beta – straight chain, used for structural components • Animals (insects) use chitin gluNAc b(1->4) gluNAc • Plants use cellulose glu b(1->4) glu • Bacteria use peptidoglycan (heteropolysaccharide b(1->4)) • Saturated hydrogen bond network • OH --- O hydrogen bonds • Weak CH --- O hydrogen bonds between sheets
Glycosylation • Proteoglycan • Heavily glycosylated (mostly sugars, little protein) • Covalently attached glycosaminoglycans (GAG) chains ex. Keratan, chondroitin sulfates O-linked to core protein • Hyaluronic acid (polysaccharide) backbone • Makes up extracellular matrix (ECM) • Together with collagen (triple amino acid helix), makes cartilage • Glycoproteins • Mostly proteins with some glycosylation • N-linked glycosylation on the outer (ECM) surface of membrane/transmembrane proteins • Indicates proper protein folding going through ER and Golgi network, used as antigens or ID markers on free proteins such as antibodies (human IgG) • Peptidoglycan • GlcNAc b(1>4) MurNAc backbone • L-Ala-Isoglu-L-Lys-D-Ala at 3’ carbon of MurNAc • E-Amino link on L-Lys to Pentaglycine bridge to Carboxyl end of D-Ala
Glycosylation • O-Linked • Gal b(1->3) GlcNAc alpha linked to Ser/Thr • N-Linked • Asn-X-Ser/Thr sequence protein side • Asn linked to beta GlcNAc • 2xMan-GlcNAc-GlcNAc-Asn sequence oligosaccharide side
Phospholipids • Glycerol backbone (three carbons each attached to OH) • 2 Nonpolar FA tails • Palmitic acid (16C) saturated • Stearic acid (18C) saturated • Oleic/Linoleic acid (16C:1 or 2 or 3) mono or poly unsaturated) • More unsaturation -> Lower Tm • Polar Head group • Phosphate(-) -> Phosphatidic acid (PA) • Phospho(-)choline(+) -> Phosphatidyl choline (PC) • Phospho(-)ethanolamine(+) -> Phosphatidylethanolamine (PE) • Phospho(-)inositol (ring) -> Phosphatidylinositol (PI) • Phospho(-)serine -> Phosphatidylserine (PS) • Phospho(-)glycerol(-) -> Phosphatidylglycerol (PG)
Membranes • Fluid Mosaic • Fluid – Constant movement • Transverse movement – rapid, often • Inversions – very very slow, rare • Both sides are different • Lipid rafts, crystalline/gel and fluid structures • Membrane proteins • Integral (lipoanchored or transmembrane) • Peripheral • Extramembraneous domains • N-linked glycoysylation on ECM side • Transmembrane proteins must have polar resides near polar head groups and nonpolar residues in hydrophobic FA tails (core
Lipids • Usually amphipathic lipids • 1 FA tail is conic, forms micelles (no aqueous interior) – called “detergents” • 2 FA tails is cylindrical, forms vesicles (bilayer forms aqueous interior) • Can be used to dislodge membrane proteins from membrane • DPPC (dipalmitoyl PC) • POPC (palmitooleyl PC)
Cell-Surface Receptor Systems • Movement of membrane proteins depend on membrane fluidity • Tyrosine autophosphorylation when two membrane proteins dimerize • Ligand binding causes conformational change->protein movement->signaling cascade