Lecture 21 Chapter 33 Nucleo4des and Nucleic Acids We have discovered the secret of life Francis Crick to patrons of The Eagle a pub in Cambridge England 1953 Francis Crick right and James Watson leK point out features of their model for the structure of DNA Outline Central Dogma review Di erences between DNA and RNA Nitrogenous bases Nucleo4des Nucleic acids DNA Composi4on of DNA Charga s Rule The DNA double helix Watson Crick Secondary structure of DNA A B Z DNA Denatura4on and Renatura4on of DNA Higher order packing of DNA in cells chromosomes and histones Nucleic acids RNA secondary and ter4ary structures of RNA Central Dogma DNA and RNA di er in the sugar component and one of the bases Alpha or Beta Base Nucleoside Base sugar Base DNA and RNA di er in the sugar component and one of the bases P Base P Base Nucleo4de Base sugar phosphate Backbones of DNA and RNA DNA 5 3 Phosphodiester bridges RNA Nitrogenous Bases Pyrimidines Structure The pyrimidine ring Found in both RNA and DNA Found in RNA only Found in DNA only Nitrogenous Bases Purines Structure The purine ring system Both are found in both RNA and DNA The Deoxyribonucleo4des Bases Adenine Guanine Thymine N glycosidic bond links N1 of pyrimidines to C1 of deoxyribose N9 of purines Cytosine The Ribonucleo4des Bases Adenine Guanine Uracyl N glycosidic bond links N1 of pyrimidines to C1 of ribose N9 of purines Cytosine Nucleotides High energy phosphoanhydride bonds Low energy phosphomonoester bond Cyclic Nucleo4des The phosphoric acid moiety is esteri ed to two of the available OH groups on the ribose ring cAMP The polymeric structure of nucleic acids Sense Strand An4sense Strand An4sense Strand Sense Strand 5 5 3 3 OH OH RNA DNA axis Chemical Proper4es of purines and pyramidines 1 UV light absorbance important for measuring DNA concentra4on 2 Nucleo4des are polypro4c acids Purines and Pyrimidines Absorp4on of UV light Pyrimidines and Purines typically strongly absorb at UV at 260 nm Nucleo4des ribo and 2 deoxyribo are polypro4c acids At pH 7 A monomeric nucleo4de nucleoside monophosphate has net charge of 2 Nucleic acids polymers of nucleo4des derive their name due to the acidic nature of the phosphate groups of their component nucleo4des 5 P end Nucleic acids have direc4onality O Reading direc4on 5 ACGU 3 5 p ACGU 3 5 pApCpGpU 3 RNA H 3 OH end Conventions in writing DNA RNA sequences Usually if you write a duplex sequence the top strand is 5 3 left to right 5 ATGTCG 3 3 TACAGC 5 Complement to top strand the bottom strand can be written as 5 CGACAT 3 or 3 TACAGC 5 forward backward Chargaff s Rules 1951 52 ds The GC or AT varies between organisms but always A T and G C Purines Pyrimidines The Double Helix Watson Crick Based on Charga s rule of DNA base composi4on X ray di rac4on data obtained by Rosalind Franklin and Maurice Wilkins James Watson and Francis Crick concluded that DNA is a complementary Double Helix Double Helix Note that the strands are an4parallel phosphate deoxyribose H bonds Watson and Crick base pairs Double Helix B form 10 4 nucleo4des per turn Axial view looking down the helix axis Double Helix The Major and Minor Grooves are lined sequence speci c hydrogen bonding groups Watson and Crick base pairs S S S deoxyribose S S Major and Minor groove of the DNA double helix Base Stacking Contribute to the Stability of the Double Helix Right handed A and B DNA Pitch 2 46 nm A form Pitch 3 4 nm B form Z DNA B DNA LeK handed helix Thinner and slimmer than B DNA DNA replica4on is Semi Conserva4ve An4sense Sense nucleo des A S A S What would conserva4ve replica4on look like A diagram of semiconserva4ve replica4on Only blue nucleo4des nucleo des without blue nucleo4des nucleo des without blue nucleo4des The resolu4on of 14N dsDNA and 15N dsDNA by density gradient centrifuga4on UV absorp4on Light DNA Heavy DNA ds double strand ss single strand Densitometric tracing Meselson Stahl experiment 1958 DNA replica4on is semiconserva ve hybrids N14 N14 N14 N15 Meselson Stahl experiment 1958 N14 N14 Parental dsDNA Denatura4on of DNA secondary structure Denaturing agents 1 Heat DNA denatura4on DNA mel4ng 2 Extremes of pH 3 Strong H bonding solutes DNA Melting Curves 1 4 Absorbance ssDNA 1 2 1 0 dsDNA Denatured DNA can renature to re form the double helix Upon cooling returning pH to neutral or when denaturants are diluted out denatured DNA will renature reanneal to re form the duplex Renatura4on is dependent on both DNA concentra4on and 4me An electron micrograph of part of the E coli genome Circular DNA from Mitochondria Relaxed dsDNA Most DNA in cells is about 5 underwound rela4ve to B form Supercoiled dsDNA Supercoiling in DNA 260 bp Underwinding Right handed DNA B DNA produces nega4ve le8 supercoils These supercoils are right handed Supercoils DNA can adopt regular structures of higher complexity Supercoiling does not mean the DNA is more or less coiled than B form DNA It means that there is a coiling super imposed on the coiling of B form DNA Double stranded circular DNA form supercoils when the two strands are underwound vely supercoiled or overwound vely supercoiled Topoisomerases are enzymes that can break one or both strands of the DNA wind them 4ghter or looser and rejoin the ends DNA Gyrase is a topoisimerase that introduces ve supercoils into DNA DNA in cells occurs in highly folded compact structures Prokaryo4c Single dsDNA molecule length 1 6 mm E Coli cell 0 002 mm length Eukaryo4c 23 pairs of ds DNA 46 DNA molecules 3 6 m total Human cell 20 m diameter Nucleus 5 m diameter Nucleosomes are complexes of DNA and histones Wrapping DNA around histones introduces nega4ve supercoils Eukaryo4c DNA is wrapped around histones to form nucleosomes 2 nm nucleosomes 146 base pair of DNA around each nucleosome core 10 nm Chroma4n DNA histone Histones are rich Lys and Arg Still more compaction is required to form condensed mitotic chromosomes Net Result Each DNA molecule has been packaged into a mitotic chromosome that is 50 000X shorter than the simple DNA helix Secondary Structures for Nucleic Acids Stem loop structures Palindromic Sequences Can Form Transient Cruciform Structures palindrome NOT the same as KAYAK Holiday junc4ons Play a role in gene4c recombina4on Single strand RNA structure Watson and Crick base pairs Non Watson and Crick base pairs rRNA ribosomal RNA 16S rRNA 2 structure 5 3 rRNA molecules have characteris4c 2 structure due to extensive intramolecular base pairing interac4ons Learning Goals Know the
View Full Document
Unlocking...