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DNA%20STRUCTURE

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IN-CLASS QUESTION: WHAT FORM OF DNA WOULD YOU EXPECT TO SEE IN DESSICATED BRINE SHRIMP EGGS? ... STABILIZES DNA AND RNA STRUCTURES ... – PowerPoint PPT presentation

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Title: DNA%20STRUCTURE


1
DNA STRUCTURE
  • STRUCTURE, FORCES AND TOPOLOGY

2
DNA GEOMETRY
  • A POLYMER OF DEOXYRIBONUCLEOTIDES
  • DOUBLE-STRANDED
  • INDIVIDUAL deoxyNUCLEOSIDE TRIPHOSPHATES ARE
    COUPLED BY PHOSPHODIESTER BONDS
  • ESTERIFICATION
  • LINK 3 CARBON OF ONE RIBOSE WITH 5 C OF ANOTHER
  • TERMINAL ENDS 5 AND 3
  • A DOUBLE HELICAL STRUCTURE
  • COMMON AXIS FOR BOTH HELICES
  • HANDEDNESS OF HELICES
  • ANTIPARALLEL RELATIONSHIP BETWEEN 2 DNA STRANDS

3
DNA GEOMETRY
  • PERIPHERY OF DNA
  • SUGAR-PHOSPHATE CHAINS
  • CORE OF DNA
  • BASES ARE STACKED IN PARALLEL FASHION
  • CHARGAFFS RULES
  • A T
  • G C
  • COMPLEMENTARY BASE-PAIRING

4
TAUTOMERIC FORMS OF BASES
  • TWO POSSIBILITIES
  • KETO (LACTAM)
  • ENOL (LACTIM)
  • PROTON SHIFTS BETWEEN TWO FORMS
  • IMPORTANT IN ORDER TO SPECIFY HYDROGEN BONDING
    RELATIONSHIPS
  • THE KETO FORM PREDOMINATES

5
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6
MAJOR AND MINOR GROOVES
  • MINOR
  • EXPOSES EDGE FROM WHICH C1 ATOMS EXTEND
  • MAJOR
  • EXPOSES OPPOSITE EDGE OF BASE PAIR
  • THE PATTERN OF H-BOND POSSIBILITIES IS MORE
    SPECIFIC AND MORE DISCRIMINATING IN THE MAJOR
    GROOVE
  • STUDY QUESTION LOCATE ALL OF THE POSSIBILITIES
    FOR H-BONDING IN THE MAJOR AND MINOR GROOVES FOR
    THE 4 POSSIBLE BASE-PAIRS

7
STRUCTURE OF THE DOUBLE HELIX
  • THREE MAJOR FORMS
  • B-DNA
  • A-DNA
  • Z-DNA
  • B-DNA IS BIOLOGICALLY THE MOST COMMON
  • RIGHT-HANDED (20 ANGSTROM (A) DIAMETER)
  • COMPLEMENTARY BASE-PAIRING (WATSON-CRICK)
  • A-T
  • G-C
  • EACH BASE PAIR HAS THE SAME WIDTH
  • 10.85 A FROM C1 TO C1
  • A-T AND G-C PAIRS ARE INTERCHANGEABLE
  • PSEUDO-DYAD AXIS OF SYMMETRY

8
GEOMETRY OF B-DNA
  • IDEAL B-DNA HAS 10 BASE PAIRS PER TURN
  • BASE THICKNESS
  • AROMATIC RINGS WITH 3.4 A THICKNESS TO RINGS
  • PITCH 10 X 3.4 34 A PER COMPLETE TURN
  • AXIS PASSES THROUGH MIDDLE OF EACH BP
  • MINOR GROOVE IS NARROW
  • MAJOR GROOVE IS WIDE
  • IN CLASS EXERCISE EXPLORE THE STRUCTURE OF
    B-DNA. PAY SPECIAL ATTENTION TO THE MAJOR, MINOR
    GROOVES

9
A-DNA
  • RIGHT-HANDED HELIX
  • WIDER AND FLATTER THAN B-DNA
  • 11.6 BP PER TURN
  • PITCH OF 34 A
  • ? AN AXIAL HOLE
  • BASE PLANES ARE TILTED 20 DEGREES WITH RESPECT
    TO HELICAL AXIS
  • HELIX AXIS PASSES ABOVE MAJOR GROOVE
  • ? DEEP MAJOR AND SHALLOW MINOR GROOVE
  • OBSERVED UNDER DEHYDRATING CONDITIONS

10
A-DNA
  • WHEN RELATIVE HUMIDITY IS 75
  • B-DNA ? A-DNA (REVERSIBLE)
  • MOST SELF-COMPLEMENTARY OLIGONUCLEO-
  • TIDES OF lt 10 bp CRYSTALLIZE IN A-DNA CONF.
  • A-DNA HAS BEEN OBSERVED IN 2 CONTEXTS
  • AT ACTIVE SITE OF DNA POLYMERASE ( 3 bp )
  • GRAM () BACTERIA UNDERGOING SPORULATION
  • SASPs INDUCE B-DNA TO ? A-DNA
  • RESISTANT TO UV-INDUCED DAMAGE
  • CROSS-LINKING OF PYRIMIDINE BASES

11
Z-DNA
  • A LEFT-HANDED HELIX
  • SEEN IN CONDITIONS OF HIGH SALT CONCENTRATIONS
  • REDUCES REPULSIONS BETWEEN CLOSEST PHOSPHATE
    GROUPS ON OPPOSITE STRANDS (8 A VS 12 A IN B-DNA)
  • IN COMPLEMENTARY POLYNUCLEOTIDES WITH ALTERNATING
    PURINES AND PYRIMIDINES
  • POLY d(GC) POLY d(GC)
  • POLY d(AC) ? POLY d(GT)
  • MIGHT ALSO BE SEEN IN DNA SEGMENTS WITH ABOVE
    CHARACTERISTICS

12
Z-DNA
  • 12 W-C BASE PAIRS PER TURN
  • A PITCH OF 44 DEGREES
  • A DEEP MINOR GROOVE
  • NO DISCERNIBLE MAJOR GROOVE
  • REVERSIBLE CHANGE FROM B-DNA TO Z-DNA IN
    LOCALIZED REGIONS MAY ACT AS A SWITCH TO
    REGULATE GENE EXPRESSION
  • ? TRANSIENT FORMATION BEHIND ACTIVELY TRAN-
  • SCRIBING RNA POLYMERASE

13
STRUCTURAL VARIANTS OF DNA
  • DEPEND UPON
  • SOLVENT COMPOSITION
  • WATER
  • IONS
  • BASE COMPOSITION
  • IN-CLASS QUESTION WHAT FORM OF DNA WOULD YOU
    EXPECT TO SEE IN DESSICATED BRINE SHRIMP EGGS?
    WHY?

14
RNA
  • UNLIKE DNA, RNA IS SYNTHESIZED AS A SINGLE STRAND
  • THERE ARE DOUBLE-STRANDED RNA STRUCTURES
  • RNA CAN FOLD BACK ON ITSELF
  • DEPENDS ON BASE SEQUENCE
  • GIVES STEM (DOUBLE-STRAND) AND LOOP
    (SINGLE-STRAND STRUCTURES)
  • DS RNA HAS AN A-LIKE CONFORMATION
  • STERIC CLASHES BETWEEN 2-OH GROUPS PREVENT THE
    B-LIKE CONFORMATION

15
HYBRID DNA-RNA STRUCTURES
  • THESE ASSUME THE A-LIKE CONFORMATION
  • USUALLY SHORT SEQUENCES
  • EXAMPLES
  • DNA SYNTHESIS IS INITIATED BY RNA PRIMERS
  • DNA IS THE TEMPLATE FOR TRANSCRIPTION TO RNA

16
FORCES THAT STABILIZE NUCLEIC ACID STRUCTURES
  • SUGAR-PHOSPHATE CHAIN CONFORMATIONS
  • BASE PAIRING
  • BASE-STACKING,HYDROPHOBIC
  • IONIC INTERACTIONS

17
SUGAR-PHOSPHATE CHAIN IS FLEXIBLE TO AN EXTENT
  • CONFORMATIONAL FLEXIBILITY IS CONSTRAINED BY
  • SIX TORSION ANGLES OF SUGAR-PHOSPHATE BACKBONE
  • TORSION ANGLES AROUND N-GLYCOSIDIC BOND
  • RIBOSE RING PUCKER

18
TORSION ANGLES
  • SIX OF THEM
  • GREATLY RESTRICTED RANGE OF ALLOWABLE VALUES
  • STERIC INTERFERENCE BETWEEN RESIDUES IN
    POLYNUCLEOTIDES
  • ELECTROSTATIC INTERACTIONS OF PHOS. GROUPS
  • A SINGLE STRAND OF DNA ASSUMES A RANDOM COIL
    CONFIGURATION

19
THE N-GLYCOSIDIC TORSION ANGLE
  • TWO POSSIBILITIES, STERICALLY
  • SYN
  • ANTI
  • PYRIMIDINES
  • ONLY ANTI IS ALLOWED
  • STERIC INTERFERENCE BETWEEN RIBOSE AND THE C2
    SUBSTITUENT OF PYRIMIDINE
  • PURINES
  • CAN BE SYN OR ANTI

20
IN MOST DOUBLE-HELICAL STRUCTURES, ALL BASES
IN ANTI FORM
21
GLYCOSIDIC TORSION ANGLES IN Z-DNA
  • ALTERNATING
  • PYRIMIDINE ANTI
  • PURINE SYN
  • WHAT HAPPENS WHEN B-DNA SWITCHES TO Z-DNA?
  • THE PURINE BASES ROTATE AROUND GLYCOSIDIC BOND
    FROM ANTI TO SYN
  • THE SUGARS ROTATE IN THE PYRIMIDINES
  • THIS MAINTAINS THE ANTI CONFORMATIONS

22
RIBOSE RING PUCKER
  • THE RING IS NOT FLAT
  • SUBSTITUENTS ARE ECLIPSED IF FLAT
  • CROWDING IS RELIEVED BY PUCKERING
  • TWO POSSIBILITIES FOR EACH OF C2 OR C3
  • ENDO OUT-OF-PLANE ATOM ON SAME SIDE OF RING AS
    C5
  • EXO DISPLACED TO OPPOSITE SIDE
  • C2 ENDO IS MOST COMMON
  • CAN ALSO SEE C3-ENDO AND C3-EXO
  • LOOK AT RELATIONSHIPS BETWEEN THE PHOSPHATES
  • IN C3 ENDO- THE PHOSPHATES ARE CLOSER THAN IN
    C2 ENDO-

23
RIBOSE RING PUCKER
  • B-DNA HAS THE C2-ENDO-FORM
  • A-DNA IS C3-ENDO
  • Z-DNA
  • PURINES ARE ALL C3-ENDO
  • PYRIMIDINES ARE ALL C2-ENDO
  • CONCLUSION THE RIBOSE PUCKER GOVERNS RELATIVE
    ORIENTATIONS OF PHOSPHATE GROUPS TO EACH SUGAR
    RESIDUE

24
IONIC INTERACTIONS
  • THE DOUBLE HELIX IS ANIONIC
  • MULTIPLE PHOSPHATE GROUPS
  • DOUBLE-STRANDED DNA HAS HIGHER ANIONIC CHARGE
    DENSITY THAT SS-DNA
  • THERE IS AN EQUILIBRIUM BETWEEN SS-DNA AND DS-DNA
    IN AQUEOUS SOLUTION
  • DS-DNA SS-DNA
  • QUESTION WHAT HAPPENS TO THE Tm OF DS-DNA AS
    CATION INCREASES? WHY?

25
IONIC INTERACTIONS
  • DIVALENT CATIONS ARE GOOD SHIELDING AGENTS
  • MONOVALENT CATIONS INTERACT NON-SPECIFICALLY
  • FOR EXAMPLE, IN AFFECTING Tm
  • DIVALENT INTERACT SPECIFICALLY
  • BIND TO PHOSPHATE GROUPS
  • MAGNESIUM (2) ION
  • STABILIZES DNA AND RNA STRUCTURES
  • ENZYMES THAT ARE INVOLVED IN RXNS WITH NUCLEIC
    ACID USUALLY REQUIRE Mg(2) IONS FOR ACTIVITY

26
BASE STACKING
  • PARTIAL OVERLAP OF PURINE AND PYRIMIDINE BASES
  • IN SOLID-STATE (CRYSTAL)
  • VANDERWAALS FORCES
  • IN AQUEOUS SOLUTION
  • MOSTLY HYDROPHOBIC FORCES
  • ENTHALPICALLY-DRIVEN
  • ENTROPICALLY-OPPOSED
  • OPPOSITE TO THAT OF PROTEINS

27
BASE-PAIRING
  • WATSON-CRICK GEOMETRY
  • THE A-T PAIRS USE ADENINES N1 AS THE H-BOND
    ACCEPTOR
  • HOOGSTEEN GEOMETRY
  • N7 IS THE ACCEPTOR
  • SEEN IN CRYSTALS OF MONOMERIC A-T BASE PAIRS
  • IN DOUBLE HELICES, W-C IS MORE STABLE
  • ALTHOUGH HOOGSTEIN IS MORE STABLE FOR A-T PAIRS,
    W-C IS MORE STABLE IN DOUBLE HELICES
  • CO-CRYSTALLIZED MONOMERIC G-C PAIRS ALWAYS FOLLOW
    W-C GEOMETRY
  • THREE H-BONDS

28
HYDROGEN BONDING
  • REQUIRED FOR SPECIFICITY OF BASE PAIRING
  • NOT VERY IMPORTANT IN DNA STABILIZATION
  • HYDROPHOBIC FORCES ARE THE MOST IMPT.

29
THE TOPOLOGY OF DNA
  • SUPERCOILING DNAS TERTIARY STRUCTURE
  • L LINKING NUMBER
  • A TOPOLOGIC INVARIANT
  • THE OF TIMES ONE DNA STRAND WINDS AROUND THE
    OTHER
  • L T W
  • T IS THE TWIST
  • THE OF COMPLETE REVOLUTIONS THAT ONE DNA STRAND
    MAKES AROUND THE DUPLEX AXIS
  • W IS THE WRITHE
  • THE OF TIMES THE DUPLEX AXIS TURNS AROUND THE
    SUPERHELICAL AXIS

30
DNA TOPOLOGY
  • THE TOPOLOGICAL PROPERTIES OF DNA HELP US TO
    EXPLAIN
  • DNA COMPACTING IN THE NUCLEUS
  • UNWINDING OF DNA AT THE REPLICATION FORK
  • FORMATION AND MAINTENANCE OF THE TRANSCRIPTION
    BUBBLE
  • MANAGING THE SUPERCOILING IN THE ADVANCING
    TRANSCRIPTION BUBBLE

31
DNA TOPOLOGY
  • AFTER COMPLETING THE 13 IN-CLASS EXERCISES, TRY
    TO ANSWER THE FOLLOWING QUESTIONS
  • (1) THE HELIX AXIS OF A CLOSED CIRCULAR DUPLEX
    DNA IS CONSTRAINED TO LIE IN A PLANE. THERE ARE
    2340 BASE PAIRS IN THIS PIECE OF DNA AND, WHEN
    CONSTRAINED TO THE PLANE, THE TWIST IS 212.
  • DETERMINE L, W AND T FOR THE CONSTRAINED
    AND UNCONSTRAINED FORM OF THIS DNA.
  • (2) A CLOSED CIRCULAR DUPLEX DNA HAS A 100 BP
    SEGMENT OF ALTERNATING C AND G RESIDUES. ON
    TRANSFER TO A SOLUTION WITH A HIGH SALT
    CONCENTRATION, THE SEGMENT MAKES A TRANSITION
    FROM THE B-FORM TO THE Z-FORM. WHAT IS THE
    ACCOMPANYING CHANGE IN L, W. AND T?
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