Midterms Doc Dumlao

• DNA


  Chemical basis of heredity and organized into genes (fundamental units of genetic info)
• Monomeric deoxynucleotide units of DNA

  • Deoxyadenylate
  • Deoxguanylate
  • Deoxycytidylate
  • Thymidylate
• Monomeric units of DNA

  Held in polymeric form by 3',5'- phosphodiester bonds constituting a single strand
• Watson, Crick, Wilkins


  • Propose early 1950s model of double-stranded (ds) DNA
  • 2 strands are held by hydrogen bonds between purine and pyrimidine bases and van der Waals and hydrophobic interactions between stacked adjacent base pairs
  • Watson-Crick base pairs: A-T, G-C base pairs
  • This common form of DNA is right-handed
  • Canonical base pair = A-T pair – held together by 2 H-bonds
  • 2 strands possess polarity and are antiparallel (5' to 3' and 3' to 5')

  
• Template Strand


  Genetic info resides in sequence of nucleotides on 1 strand, strand of DNA that is copied/transcribed during RNA synthesis, non-coding strand
• Coding Strand


  Matches the sequence of RNA transcript (U instead of T)
• DNA


  • Contains a major and minor groove, winding along the molecule parallel to phosphodiester backbones
  • Proteins often interact with exposed atoms of nucleotides via hydrophobic and ionic interactions
  • Binding occurs without disrupting the base pairing of double-helix
• The denaturation of DNA is used to analyze its structure
• Deoxynucleotide


  • 3 bonds formed by hydrogen atoms bonded to electronegative N or O atoms hold the deoxyguanosine nucleotide to deoxycytidine nucleotide
  • 4 DNA nucleotide are flat and planar molecules
  • Atoms within the aromatic, heterocyclic bases are highly polarizable and coupled with many atoms within the bases contain partial charges, allowing stack bases to form van der Waals and electrostatic interactions (base-stacking forces/interactions)
  • G-C rich DNA sequence – more resistant to denaturation or strand separation or melting
• DNA denaturation


  • Can be done by increasing temperature, decreasing solution salt concentrations, or adding chaotropic agents (forms competing H-bonds with individual deoxynucleotide bases)
  • Denaturation of DNA in experiments by combination of all 3 treatments
  • Hyperchromicity of denaturation – increase in optical absorbance in UV light spectrum of purine and pyrimidine bases of each strand
  • On denaturation, DNA lose their viscosity
  • DNA denaturation and renaturation – occurs naturally during the processes of DNA replication, DNA recombination, DNA repair, and DNA replication
• Renaturation of DNA


  • Requires precise base pair matching
  • Rate of strand reassociation depends on concentration of complementary strand
  • At given temperature and salt concentration, a particular nucleic strand will associate tightly only with its complementary strand, renaturation is highly specific
• DNA


  • Exists in relaxed and supercoiled forms
  • Negative supercoils – formed when molecule is twisted in the direction opposite from clockwise turns of right-handed double helix found in B-DNA, such DNA is said to be underwound, energy required is stored in supercoils, strand separation – prerequisite for DNA replication and transcription
  • Topoisomerase – enzymes that catalyze topologic changes of DNA, can relax or insert supercoils using ATP as energy source, homologs exists in all organisms and an important target for cancer chemotherapy

  
• DNA


  • Provides a template for replication and transcription
  • 2 newly formed daughter dsDNA molecules each contain 1 strand (complementary) from parent dsDNA, then sorted between 2 daughter cells during mitosis, each daughter cell contains DNA molecules with information identical to that which the parent possessed
• RNA


  Chemical nature differs from DNA: Sugar moiety (RNA – ribose, DNA – deoxyribose), Pyrimidine components (RNA – Uracil, DNA – Thymine), Strand (RNA – single, DNA – double), Content (RNA – guanine does not necessarily equal to cytosine), Alkali lability (RNA – hydrolyzed by alkali to 2', 3' cyclic diesters of mononucleotides)
• RNA species involved in protein synthesis

  • mRNA – templates for protein synthesis
  • rRNA – structural role; formation and function of ribosomes
  • tRNA – serves as adapter molecules for translation of RNA into specific polymerized amino acids
  • RNA enzymes/ Ribozymes – intrinsic catalytic activity; often involves cleavage of nucleic acid
  • snRNA – do not directly involved in protein synthesis but play pivotal roles in RNA processing (mRNA)
• mRNA


  • Most heterogenous in abundance, size, and stability
  • Contain internal N6-methyladenine and other 2′-O-ribose-methylated nucleotides
  • 5′ terminus is "capped" by a 7-methylguanosine triphosphate that is linked to an adjacent 2′-O-methyl ribonucleoside at its 5′-hydroxyl through the three phosphates
  • Must be formed by processing from precursor or pre-mRNA before entering the cytoplasm
• tRNA


  • Primary structure (nucleotide sequence) allows extensive folding and intrastrand complementarity to generate secondary structure that appears like cloverleaf
  • At least 20 species of tRNA molecules in every cell
  • Also generated by nuclear processing of precursor molecule
• rRNA


  • Ribosome – cytoplasmic nucleoprotein structure that acts as machine for synthesis of proteins from mRNA
  • mRNA and tRNA interact to translate info transcribed from gene during mRNA synthesis into specific protein
  • Ribosomes associated with single mRNA to form an assembly called polysome
  • Mammalian ribosome – contains 2 major nucleotide protein subunits: 60S (5S rRNA, 5.8S rRNA, 28S rRNA; >50 polypeptides) and 40S (18S rRNA and 30 polypeptide chains)
• sRNA


  • Highly conserved small RNA found in eukaryotic cells, 20-1000 nucleotides in 100,000 to 1M copies per cell
• snRNA


  • Subset of sRNA, involved in rRNA and mRNA processing and gene regulation
  • U1, U2, U4, U5, U6 = involved in mRNA splicing (nuclear process whereby introns are removed from mRNA to generate functional cytoplasmic mRNA)
  • U7 = involved in production of correct 3' ends of histone mRNA (lacks poly(A) tail)
  • 7SK RNA = associated with several proteins to form ribonucleic complex called P-TERFb (modulates mRNA gene transcription elongation by RNA pol II)
• Classes of RNA


  • mRNA
  • tRNA
  • rRNA
  • sRNA
  • snRNA
• mRNA


  • Most heterogenous in abundance, size, and stability
  • Contains internal N6-methyladenine and other 2′-O-ribose-methylated nucleotides
  • 5′ terminus is "capped" by a 7-methylguanosine triphosphate that is linked to an adjacent 2′-O-methyl ribonucleoside at its 5′-hydroxyl through the three phosphates
  • Must be formed by processing from precursor or pre-mRNA before entering the cytoplasm
• tRNA


  • Primary structure (nucleotide sequence) allows extensive folding and intrastrand complementarity to generate secondary structure that appears like cloverleaf
  • At least 20 species of tRNA molecules in every cell
  • Also generated by nuclear processing of precursor molecule
• Ribosome


  • Cytoplasmic nucleoprotein structure that acts as machine for synthesis of proteins from mRNA
  • mRNA and tRNA interact to translate info transcribed from gene during mRNA synthesis into specific protein
  • Associated with single mRNA to form an assembly called polysome
  • Mammalian ribosome contains 2 major nucleotide protein subunits: 60S and 5S rRNA, 5.8S rRNA, 28S rRNA; >50 polypeptides
• sRNA


  • Highly conserved small RNA found in eukaryotic cells
  • 20-1000 nucleotides in 100,000 to 1M copies per cell
• snRNA


  • Highly conserved small RNA found in eukaryotic cells
  • Subset of sRNA
  • Involved in rRNA and mRNA processing and gene regulation
  • U1, U2, U4, U5, U6 involved in mRNA splicing
  • U7 involved in production of correct 3' ends of histone mRNA
  • 7SK RNA associated with several proteins to form ribonucleic complex called P-TERFb (modulates mRNA gene transcription elongation by RNA pol II)
• Specific nucleases digest nucleic acids
• Deoxyribonucleases exhibit specificity for DNA
• Ribonucleases specifically hydrolyze RNA
• Endonucleases produce either 3′-hydroxyl and 5′-phosphoryl terminals or, 5′-hydroxyl and 3′-phosphoryl terminals
• Exonucleases are capable of hydrolyzing nucleotide only when it is present at terminal of a molecule
• Chromatin


  Chromosomal material in the nuclei of cells of eukaryotic organisms, consisting of double-stranded DNA (dsDNA) molecules, histones, non-histones, and small quantity of RNA
• Histones


  • Most abundant chromatin proteins
  • Nucleosomes contain 4 types: H2A, H2B, H3, H4
• Centromere base pair is AT, Telomere base pair is TG
• At least 30% of the human genome consists of repetitive sequences
• DNA in eukaryotic genome can be divided into nonrepetitive (unique-sequence) and repetitive-sequence DNA
• DNA includes single copy of genes that code for proteins
• Polymerase Chain Reaction (PCR)


  • Detects heritable traits
  • Rapidly screens for microsatellite polymorphism
• Indels


  Insertions or deletions, often result in mutation
• Jumping DNA/Jumping Genes


  Can carry flanking regions of DNA, profoundly affect evolution