Chapter 16 The Molecular Basis of Inheritance. Fig. 16-1 In 1953, James Watson and Francis Crick introduced an elegant double-helical model for the structure.

Chapter 16

The Molecular Basis of Inheritance

Fig. 16-1

•In 1953, James Watson and Francis Crick introduced an elegant double-helical model for the structure of deoxyribonucleic acid, or DNA

Fig. 16-2

Living S cells (control) Pathogenic

Living R cells (control)

Heat-killed S cells (control)

Mixture of heat-killed S cells and living R cells

Mouse diesMouse dies Mouse healthy Mouse healthy

Living S cells

RESULTS

EXPERIMENT Frederick Griffith in 1928

Transformation,

now defined as a change in genotype and phenotype due to assimilation of foreign DNA

•In 1944, Oswald Avery, Maclyn McCarty, and Colin MacLeod announced that the transforming substance was DNA

Hershey and Chase

• 1952, studying T2 virus infecting Escherichia coli– Bacteriophage or phage

• Phage coat made entirely of protein• DNA found inside capsid

Fig. 16-4-1

EXPERIMENT

Phage

DNA

Bacterial cell

Radioactive protein

Radioactive DNA

Batch 1: radioactive sulfur (35S)

Batch 2: radioactive phosphorus (32P)

Fig. 16-4-2

EXPERIMENT

Phage

DNA

Bacterial cell

Radioactive protein

Radioactive DNA



Empty protein shell

Phage DNA

Fig. 16-4-3

EXPERIMENT

Phage

DNA

Bacterial cell

Radioactive protein

Radioactive DNA



Empty protein shell

Phage DNA

Centrifuge

Centrifuge

Pellet

Pellet (bacterial cells and contents)

Radioactivity (phage protein) in liquid

Radioactivity (phage DNA) in pellet

Chargaff’s rules

• It was known that DNA is a polymer of nucleotides, each consisting of a nitrogenous base, a sugar, and a phosphate group

• In 1950, Erwin Chargaff reported that DNA composition varies from one species to the next

• Chargaff’s rules state that in any species there is an equal number of A and T bases, and an equal number of G and C bases

Fig. 16-5Sugar–phosphate

backbone

5 end

Nitrogenous

bases

Thymine (T)

Adenine (A)

Cytosine (C)

Guanine (G)

DNA nucleotide

Sugar (deoxyribose)

3 end

Phosphate

Building a Structural Model of DNA: Scientific Inquiry

• After most biologists became convinced that DNA was the genetic material, the challenge was to determine how its structure accounts for its role

• Maurice Wilkins and Rosalind Franklin were using a technique called X-ray crystallography to study molecular structure

• Franklin produced a picture of the DNA molecule using this technique

Fig. 16-6

(a) Rosalind Franklin (b) Franklin’s X-ray diffraction photograph of DNA

Fig. 16-7

(c) Space-filling model

Hydrogen bond 3 end

5 end

3.4 nm

0.34 nm

3 end

5 end

(b) Partial chemical structure(a) Key features of DNA structure

1 nm

• DNA is– Double stranded– Helical– Sugar-phosphate

backbone– Bases on the

inside– Stabilized by

hydrogen bonding– Base pairs with

specific pairing

• AT/GC or Chargoff’s rule– A pairs with T

– G pairs with C

• Keeps with consistent

• 10 base pairs per turn

• 2 DNA strands are complementary– 5’ – GCGGATTT – 3’

– 3’ – CGCCTAAA – 5’

• 2 strands are antiparallel– One strand 5’ to 3’

– Other stand 3’ to 5’

•two antiparallel sugar-phosphate backbones, with the nitrogenous bases paired in the molecule’s interior

• Space-filling model shows grooves– Major groove

• Where proteins bind

– Minor groove

Fig. 16-7b

Replication

• 3 different models for DNA replication proposed in late 1950s– Semiconservative– Conservative– Dispersive

• Newly made strands are daughter strands

• Original strands are parental strands

Fig. 16-10

Parent cellFirst replication

Second replication

(a) Conservative model

(b) Semiconserva- tive model

(c) Dispersive model

Fig. 16-11EXPERIMENT

RESULTS

CONCLUSION

1 2

43

Conservative model

Semiconservative model

Dispersive model

Bacteria cultured in medium containing 15N

Bacteria transferred to medium containing 14N

DNA sample centrifuged after 20 min (after first application)

DNA sample centrifuged after 40 min (after second replication) More

dense

Less dense

Second replicationFirst replication

Fig. 16-11a

EXPERIMENT

RESULTS

1

3

2

4

Bacteria cultured in medium containing 15N

Bacteria transferred to medium containing 14N

DNA sample centrifuged after 20 min (after first application)

DNA sample centrifuged after 20 min (after second replication)

Less dense

More dense

a rare heavy form (15N)

a common light form (14N)

Fig. 16-11b

CONCLUSION

First replication Second replication

Conservative model

Semiconservative model

Dispersive model

DNA Replication: A Closer Look

• The copying of DNA is remarkable in its speed and accuracy

• More than a dozen enzymes and other proteins participate in DNA replication

• Origin of replication– Site of start point for replication

• Bidirectional replication– Replication proceeds outward in opposite directions

• Bacteria have a single origin

Eukaryotes require multiple origins

Fig. 16-12Origin of replication Parental (template) strand

Daughter (new) strand

Replication fork

Replication bubble

Two daughter DNA molecules

(a) Origins of replication in E. coli

Origin of replication Double-stranded DNA molecule

Parental (template) strandDaughter (new) strand

Bubble Replication fork


(b) Origins of replication in eukaryotes

0.5 µm

0.25 µm

Double-strandedDNA molecule

Fig. 16-12a

Origin of replication Parental (template) strand

Daughter (new) strand

Replication fork

Replication bubble

Double-stranded DNA molecule


(a) Origins of replication in E. coli

0.5 µm

Fig. 16-12b

0.25 µm

Origin of replication Double-stranded DNA molecule

Parental (template) strandDaughter (new) strand

Bubble Replication fork


(b) Origins of replication in eukaryotes

Topoisomerase

Helicase

PrimaseSingle-strand binding proteins

RNA primer

5 5

53

3

3

DNA helicaseBinds to DNA and travels 5’ to 3’ using ATP to separate strand and move fork forward

DNA topoisomerase: Relives additional coiling ahead of replication forkSingle-strand binding proteins: Keep parental strands open to act as templates

DNA polymerasescannot initiate synthesis of a polynucleotide; they can only add nucleotides to the 3 endPrimerase:The initial nucleotide strand is a short RNA primer

•The primer is short (5–10 nucleotides long), and the 3 end serves as the starting point for the new DNA strand

• At the end of each replication bubble is a replication fork, a Y-shaped region where new DNA strands are elongating

• Helicases are enzymes that untwist the double helix at the replication forks

• Single-strand binding protein binds to and stabilizes single-stranded DNA until it can be used as a template

• Topoisomerase corrects “overwinding” ahead of replication forks by breaking, swiveling, and rejoining DNA strands

Fig. 16-14

A

C

T

G

G

G

GC

C C

C

C

A

A

AT

T

T

New strand 5 end

Template strand 3 end 5 end 3 end

3 end

5 end5 end

3 end

Base

Sugar

Phosphate

Nucleoside triphosphate

Pyrophosphate

DNA polymerase

DNA polymerases

• Enzymes called DNA polymerases catalyze the elongation of new DNA at a replication fork

• Most DNA polymerases require a primer and a DNA template strand

• The rate of elongation is about 500 nucleotides per second in bacteria and 50 per second in human cells

Fig. 16-15

Leading strand

Overview

Origin of replicationLagging strand

Leading strandLagging strand

Primer

Overall directions of replication

Origin of replication

RNA primer

“Sliding clamp”

DNA poll IIIParental DNA

5

3

3

3

3

5

5

5

5

5

Fig. 16-15a

Overview

Leading strand

Leading strandLagging strand

Lagging strand


Primer


Fig. 16-15b


RNA primer

“Sliding clamp”

DNA pol IIIParental DNA

3

5

5

5

5

5

5

3

3

3

Fig. 16-16Overview


Leading strand

Leading strand

Lagging strand

Lagging strand


Template strand

RNA primer

Okazaki fragment

Overall direction of replication

12

3

2

1

1

1

1

2

2

51

3

3

3

3

3

3

3

3

3

5

5

5

5

5

5

5

5

5

5

53

3

Fig. 16-16a

Overview


Leading strand

Leading strand

Lagging strand

Lagging strand


12

Fig. 16-16b1

Template strand

5

53

3

Fig. 16-16b2

Template strand

5

53

3

RNA primer 3 5

5

3

1

Fig. 16-16b3

Template strand

5

53

3

RNA primer 3 5

5

3

1

1

3

35

5

Okazaki fragment

Fig. 16-16b4

Template strand

5

53

3

RNA primer 3 5

5

3

1

1

3

35

5

Okazaki fragment

12

3

3

5

5

Fig. 16-16b5

Template strand

5

53

3

RNA primer 3 5

5

3

1

1

3

35

5

Okazaki fragment

12

3

3

5

5

12

3

3

5

5

Fig. 16-16b6

Template strand

5

53

3

RNA primer 3 5

5

3

1

1

3

35

5

Okazaki fragment

12

3

3

5

5

12

3

3

5

5

12

5

5

3

3

Overall direction of replication

Fig. 16-17

OverviewOrigin of replication

Leading strand

Leading strand

Lagging strand

Lagging strandOverall directions

of replication

Leading strand

Lagging strand

Helicase

Parental DNA

DNA pol III

Primer Primase

DNA ligase

DNA pol III

DNA pol I

Single-strand binding protein

5

3

5

5

5

5

3

3

3

313 2

4

Table 16-1

Proofreading and Repairing DNA

• DNA polymerases proofread newly made DNA, replacing any incorrect nucleotides

• In mismatch repair of DNA, repair enzymes correct errors in base pairing

• DNA can be damaged by chemicals, radioactive emissions, X-rays, UV light, and certain molecules (in cigarette smoke for example)

• In nucleotide excision repair, a nuclease cuts out and replaces damaged stretches of DNA

Fig. 16-18

Nuclease

DNA polymerase

DNA ligase

Replicating the Ends of DNA Molecules

• Limitations of DNA polymerase create problems for the linear DNA of eukaryotic chromosomes

• The usual replication machinery provides no way to complete the 5 ends, so repeated rounds of replication produce shorter DNA molecules

Fig. 16-19

Ends of parental DNA strands

Leading strand

Lagging strand

Lagging strand

Last fragment Previous fragment

Parental strand

RNA primer

Removal of primers and replacement with DNA where a 3 end is available

Second round of replication

New leading strand

New lagging strand

Further rounds of replication

Shorter and shorter daughter molecules

5

3

3

3

3

3

5

5

5

5

Telomeres and aging

• Body cells have a predetermined life span

• Skin sample grown in a dish will double a finite number of times– Infants, about 80 times– Older person, 10 to 20 times

• Senescent cells have lost the capacity to divide

• Progressive shortening of telomeres correlated with cellular senescence

• Telomerase present in germ-line cells and in rapidly dividing somatic cells

• Telomerase function reduces with age

• Inserting a highly active telomerase gene into cells in the lab causes them to continue to divide

Telomeres and cancer

• When cells become cancerous they divide uncontrollably

• In 90% of all types of human cancers, telomerase is found at high levels

• Prevents telomere shortening and may play a role in continued growth of cancer cells

• Mechanism unknown

Fig. 16-20

1 µm

Fig. 16-21a

DNA double helix (2 nm in diameter)

Nucleosome(10 nm in diameter)

Histones Histone tailH1

DNA, the double helix Histones Nucleosomes, or “beads on a string” (10-nm fiber)

Concept 16.3 A chromosome consists of a DNA molecule packed together with proteins

Fig. 16-21b

30-nm fiber

Chromatid (700 nm)

Loops Scaffold

300-nm fiber

Replicated chromosome (1,400 nm)

30-nm fiber Looped domains (300-nm fiber)

Metaphase chromosome

• Chromatin is organized into fibers

• 10-nm fiber– DNA winds around histones to form nucleosome

“beads”– Nucleosomes are strung together like beads on a

string by linker DNA

• 30-nm fiber– Interactions between nucleosomes cause the thin

fiber to coil or fold into this thicker fiber

Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings

• 300-nm fiber– The 30-nm fiber forms looped domains that attach

to proteins

• Metaphase chromosome– The looped domains coil further– The width of a chromatid is 700 nm


• Most chromatin is loosely packed in the nucleus during interphase and condenses prior to mitosis

• Loosely packed chromatin is called euchromatin

• During interphase a few regions of chromatin (centromeres and telomeres) are highly condensed into heterochromatin

• Dense packing of the heterochromatin makes it difficult for the cell to express genetic information coded in these regions


• Histones can undergo chemical modifications that result in changes in chromatin organization– For example, phosphorylation of a specific amino

acid on a histone tail affects chromosomal behavior during meiosis


Fig. 16-22

RESULTS

Condensin and DNA (yellow)

Outline of nucleus

Condensin (green)

DNA (red at periphery)

Normal cell nucleus Mutant cell nucleus

You should now be able to:

1. Describe the contributions of the following people: Griffith; Avery, McCary, and MacLeod; Hershey and Chase; Chargaff; Watson and Crick; Franklin; Meselson and Stahl

2. Describe the structure of DNA

3. Describe the process of DNA replication; include the following terms: antiparallel structure, DNA polymerase, leading strand, lagging strand, Okazaki fragments, DNA ligase, primer, primase, helicase, topoisomerase, single-strand binding proteins

4. Describe the function of telomeres

5. Compare a bacterial chromosome and a eukaryotic chromosome

Fig. 16-UN5

Chapter 16 The Molecular Basis of Inheritance. Fig. 16-1 In 1953, James Watson and Francis Crick introduced an elegant double-helical model for the structure.

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