Chapter 2 & 3: DNA Structure and

Replication

Ms. Gaynor

Honors Genetics

DNA and Its Structure (Part 2)

•From 1953

Recall…•DNA and RNA are nucleic acids

•An important macromolecule in organisms that stores and carries genetic information

What is the Double Helix?

•Shape of DNA•Looks like a twisted ladder

•2 coils are twisted around each other

•Double means 2•Helix means coil

The Structure of DNA • Made out of nucleotides

•Includes a phosphate group, nitrogenous base and 5-carbon pentose sugar

Nucleotide Structure

1 “link” in a DNA chain

A Polynucleotide

• MANY nucleotides (“links”) bonded togetherDNA has a

overall negative

charge b/c of the PO4

-3 (phosphate

group)

The Structure of DNA Backbone = alternating P’s and sugar

•Held together by COVALENT bonds (strong)•Inside of DNA molecule = nitrogen base pairs

•Held together by HYDROGEN bonds (weaker)

Backbone

• Phosphodiester Bond

–The covalent that holds together the backbone

–Found between P & deoxyribose sugar

–STRONG!!!

Major Groove

Minor Groove

DNA is antiparallel• Antiparallel means that the

1st strand runs in a 5’ 3’ direction and the 2nd 3’ 5’ direction – THEY RUN IN OPPOSITE

or ANTIPARALLEL DIRECTIONS

• P end is 5’ end (think: “fa” sound)

• -OH on deoxyribose sugar is 3’ end– 5’ and 3’ refers to the carbon

# on the pentose sugar that P or OH is attached to

DNA in Cells• 2 broad categories of cells1. Eukaryotic cells: have nucleus

with DNA– DNA is contained in

structure called a chromosome

– Chromosomes are a LINEAR (line) shape with ENDS called telomeres (protective “caps”)

2. Prokaryotic cells: no nucleus (nucleoid region instead) which contains DNA– DNA is a CIRCULAR shaped

chromosome without ENDS (no telomeres)

DNA Bonding• Purines (small word, big base)

– Adenine– Guanine

• Pyrimidines– (big word, small base)

– Cytosine

– Thymine

• Chargaff’s rules– A=T, C=G– Hydrogen BondsHydrogen Bonds attractions between the

stacked pairs; WEAK bonds

Why Does a Purine Always Bind with A Pyrimidine?

DNA Double Helix• http://www.sumanasinc.com/webcontent/animations/content/DNA_structure.html• Watson & Crick said that…

– strands are complementary; nucleotides line up on template according to base pair rules (Chargaff’s rules)

• A to T and C to G

•LET’S PRACTICE…– Template: 5’AATCGCTATAC3’

– Complementary strand: 3’ TTAGCGATATG5’

DNA Replication(Part 3A-initiation)

DNA Replication• DNA Replication =

DNA DNA– Parent DNA makes 2

exact copies of DNA– Why??

•Occurs in Cell Cycle before MITOSIS so each new cell can have its own FULL copy of DNA

http://www.sumanasinc.com/webcontent/animations/content/meselson.html

Models of DNA Replication

DNA Replication• How does it occur?

• Matthew Meselson & Frank Stahl

– Discovered replication is semiconservative– PROCEDURE varying densities of radioactive nitrogen (Nitrogen is in DNA)

Meselson & Stahl Experiment**DNA is semiconservative

http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter14/animations.html

DNA Replication: a closer look

http://henge.bio.miami.edu/mallery/movies/replication.mov

DNA Replication Steps:• Initiation

– involves assembly of replication fork (bubble) at origin of replication • sequence of DNA found at a specific

site • Elongation

– Parental strands unwind and daughter strands are synthesized.

– the addition of bases by proteins• Termination:

– the duplicated chromosomes separate from each other. Now, there are 2 IDENTICAL copies of DNA.

Segments of single-stranded DNA are called template strands.

Copied strand is called the complement strand (think “c” for copy)

BEGINNING OF DNA REPLICATION(INITIATION)

• Gyrase (type of topoisomerase) – relaxes the supercoiled DNA.

• DNA helicase (think “helix”) – binds to the DNA at the replication fork – untwist (“unzips”) DNA using energy from ATP– Breaks hydrogen bonds between base pairs

• Single-stranded DNA-binding proteins (SSBP) – stabilize the single-stranded template DNA

during the process so they don’t bond back together.

base pairs

5’

5’

3’

3’

Supercoiled DNA relaxed by gyrase & unwound by helicase

Helicase

ATPSSB Proteins

http://media.pearsoncmg.com/bc/bc_campbell_biology_7/media/interactivemedia/activities/load.html?16&F

SSB Proteins

Gyrase

DNA Replication (Elongation)After SSBP’s bind to each template…• RNA Primase binds to helicase

– primase is required for DNA synthesis– Like a “key” for a car ignition – makes a short RNA primers

• Short pieces of RNA needed for DNA synthesis• DNA polymerase

– adds nucleotides to RNA primer makes POLYNUCLEOTIDES (1st function)

– After all nucleotides are added to compliment strand…• RNA primer is removed and replaced with

DNA by DNA polymerase (2nd function)• DNA ligase

– “seals” the gaps in DNA – Connects DNA pieces by making phosphodiester bonds

DNA Polymerase

Leading strand

base pairs

5’

5’

3’

3’

Supercoiled DNA relaxed by gyrase & unwound by helicase + proteins:

Helicase

ATP

SSB Proteins

RNA Primer

primase

2DNA Polymerase

1

RNA primer replaced by DNA Polymerase & gap is sealed by ligase

http://media.pearsoncmg.com/bc/bc_campbell_biology_7/media/interactivemedia/activities/load.html?16&F

Gyrase

ElongationAntiparallel nature: • Sugar (3’end)/phosphate

(5’ end) backbone runs in opposite directions – one strand runs 5’ 3’,– other runs 3’ 5’

• DNA polymerase only adds nucleotides at the free 3’ end of NEW STRAND forming new DNA strands in the 5’ 3’ direction

only!!!

Elongation (con’t)

• Leading (daughter) strand– NEW strand made toward the

replication fork (only in 5’ 3’ direction from the 3’ 5’ master strand

– Needs ONE (1) RNA primer made by Primase

– This new leading strand is made CONTINOUSLY

Elongation (con’t)Lagging (daughter) strand• NEW strand synthesis away from

replication fork

• Replicate DISCONTINUOUSLY – Creates Okazaki fragments

• Short pieces of DNA

– Okazaki fragments joined by DNA ligase

• “Stitches” fragments together

– Needs MANY RNA primer made by Primase

•

3

DNA Polymerase

5’ 3

’

Leading strand

base pairs

5’

5’

3’

3’

Supercoiled DNA relaxed by gyrase & unwound by helicase + proteins:

Helicase

ATP

SSB Proteins

RNA Primer

primase

2DNA Polymerase

Lagging strand

Okazaki Fragments

1

RNA primer replaced by DNA Polymerase & gap is sealed by DNA ligase

http://media.pearsoncmg.com/bc/bc_campbell_biology_7/media/interactivemedia/activities/load.html?16&F

Gyrase

DNA Replication:Elongation

http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter14/animations.html

DNA Replication(Part 3C-termination)

Termination (Telomeres)• Telomeres

– Short repeats of “G” base found at END of LINEAR chromosomes in eukaryotes

– protect ends of linear chromosomes

– The repeated sequence of GGGTTA make up the human telomeres.

• Telomerase is the enzyme that makes telomeres.

Telomeres, Aging & Cancer

• Telomeres get shorter as cell divides leads to aging???

• Most cancers come from body cells.– Cancers cell have ability to divide

indefinitely.– Normal cells limited to ~50-75

divisions stop making telomerase.– 85–90% cancer cells continue to

make high levels of telomerase & are able to prevent further shortening of their telomeres.• Leads to “immortality”

Mistakes Made during DNA Replication

• Mutation– Change in DNA (genetic material)

•Frameshift(s)

–extra or missing base(s).

•Substitutions

–when the wrong nucleotide is incorporated (mismatch mutation).

•Deletions–Nucleotides are deleted shortening the

DNA

DNA RepairErrors occur 1/10 billion

nucleotides(Humans have 3 billion base pairs in their DNA)

• Mismatch repair– DNA polymerase (yes…it’s 3rd function)

•“Proofread” new DNA– Like the “delete” key on computer

• Excision (“cut out”) repair– Nuclease

DNA Repair