LEHNINGER PRINCIPLES OF BIOCHEMISTRY

Fifth Edition

David L. Nelson and Michael M. Cox

© 2008 W. H. Freeman and Company

CHAPTER 24Information Pathways:Genes and

Chromosomes

Central Dogma: introduced by Francis Crickthe general pathways of information flow via replication, transcription, and

translation

1953 년 James Watson Francis Crick : DNA double helix 구조규명

Chapter 24. Genes and Chromosomes1. Chromosome 의 구성성분2. DNA supercoiling3. Chromosome 의 구조

• Genes: the fundamental unit of information in living systems– 생물학적 기능을 갖는 산물을 만드는데 필요한 정보를 암호화하는 DNA 의 일부

• Chromosomes: DNA 가 packaged 되어있는 구조 , 염색체 , contains thousands of genes and intergenic DNA

• Chromatin( 염색질 ): the entire complex of a eukaryotic chromosome, including DNA, chromosomal proteins and RNA

• Chromatid( 염색분체 ): one of the two daughter strands of a replicated chromosome( 복제결과 만들어진 자매염색 분체 )

• Genome: 생물의 최소한의 유전자군 ( 群 ) 을 가진 염색체의 한 세트 , 한 세포에 함유되는 DNA 의 총체

George Beadle, Edward Tatum: proposed a molecular definition of a gene in 1940 : one gene-one enzyme one

gene- one protein

FIGURE 24-2 Colinearity of the coding nucleotide sequences of DNA and mRNA and the amino acid sequence of a polypeptide chain.

1. Chromosomal elements• Genes are segments of DNA that code for polypeptide

chains and RNAs– Phenotype( 표현형 ): visible property (ex. Eye color)– George Beadle, Edward Tatum: proposed a molecular definition of

a gene in 1940– One gene-one enzyme– One gene-one protein– One gene-one polypeptide, tRNA, rRNA– Regulatory sequences provide signals of the beginning, the end of

gene, transcription, initiation points for replication, recombination

• E. coli– Completely sequenced, a circular DNA– Base pairs: 4,639,221 bp– Code: 4,300 genes for proteins, 115 genes for stable RNAcf) Human: 3.2 billion bp, 30,000 to 35,000 gene, 24 different

chromosomes

The size and sequence structure of DNA molecules

• Viral DNA molecules are relatively small.– Less genetic information– Genome (a single RNA or DNA) and protein coat– Retrovirus and DNA virus– Replicative forms: become circular and double-stranded

• Bacteria contain chromosomes and extrachromosomal DNA– More DNA than virus (100 times much E. coli vs bacteriophage )– A single double-stranded circular DNA– More tightly compacted tertiary structure than viral DNA– Circular DNA chromosome in the nucleoid– Extrachromosomal elements( 염색체외 DNA): plasmids free in cytosol

• 1,000 to 100,000 bp long• Carry generic information• Undergo replication (self propagation)• Antibiotic resistant cell ( 항생제의 남용 )• Cloning, recombinant DNA

FIGURE 24-1 Bacteriophage T2 protein coat surrounded by its single, linear molecule of DNA.

Bacteriophage T2 protein coat surrounded by single, linear molecule of DNA

2μm

FIGURE 24-4 DNA from a lysed E. coli cell.

Chromosome 길이 : 1.7mm

The size and sequence structure of DNA molecules

• Eukaryotic cells contain more DNA than do prokaryotes.– More DNA than E.coli (yeast (2.6 배 ), fruit fly (35 배 ), human (700 배 ))– 50 genes/mm of human DNA vs. 2,500 genes/mm of E.coli DNA– 각세포는 2 m DNA x 1014 cells( 사람 ) = 2 x 1011 km

• 지구의 둘레 : 4 x 104 km, 태양까지의 거리 : 1.5 x 108 km– DNA compaction– Diploid (2n) number depends upon the species

• Human– 24 different types of chromosomes (22 matching pairs plus X and Y

set chromosomes) 25 배 이상의 길이차이 , 그러나 gene 의 수는 별차이 없음

• Organelles of eukaryotic cells also contain DNA.– Mitochondria and chloroplasts– Less than 0.1% of cell’s DNA in typical somatic cells– Much smaller than nuclear chromosomes (16,569 bp in human mtDNA)– Circular duplex, tRNA, rRNA, mitochondrial proteins– cpDNA(Chloroplast DNA): circular duplex, 120,000 to 160,000bp

FIGURE 24-5a Eukaryotic chromosomes.. FIGURE 24-5b Eukaryotic chromosomes.

FIGURE 24-6 A dividing mitochondrion.

Eukaryotic chromosomes are very complex• Intervening sequences (introns): nontranslated DNA segments in

genes (30% of human genome)– Exons: the coding segments (1.5%)– Few prokaryotic genes contain introns.– In higher eukaryotes: more introns than exons

• Moderately repetitive sequences (45% of human genome)– Derived from transposable elements (few hundreds to several thousands bp long)– Transposon: molecular parasites in host genome– Major role in human evolution: redistribution of other genomic sequences

• Highly repetitive sequences (3% of human genome)– Simple-sequenced DNA, simple sequence repeat (SSR)– Less than 10 bp long, millions times repeated per cell

• Satellites DNA: – Unusual base composition cause to migrate as satellite bands– Simple sequence DNA does not encode protein or RNA, associated with 2 important

structures– Centromere and Telomere– Artificial chromosome: centromere, telomeres, and DNA replication sequences

FIGURE 24-7 Introns in two eukaryotic genes.

Intervening sequences, intronCoding sequences, Exon

Type of sequences in human genome

Long interspersed elements6 to 8 kbpA few genes to catalyze transposition

Short interspersed elements100 to 300 bpAlu element (major)

1.5~11 kbpEvolutionally related to the retroviruses

Genes for proteins

Segmental duplicationAppear more than once

Simple-sequence repeats

Genes encoding RNAsRemnants of transposons?Unidentified: evolutionarily altered

FIGURE 24-8 Types of sequences in the human genome.

Centromere: functions during cell division as an attachment points for proteins that link the chromosomes to the mitotic spindle

yeast: 130 bp, rich in A=Thigher eukaryotes: longer, consists of simple-sequence DNATelomere: sequences at the ends of eukaryotic chromosomes that help stabilize the

chromosome•5’(TxGy)n, 3’(AxCy)n: ranges for x, y: 1 ~ 4, n: 20 ~ 100, 1500 in mammals•Telomerase: add repeated telomeric sequences•Linear DNA

Human artificial chromosomes (HACs) are being developed for somatic gene therapy

FIGURE 24-9 Important structural elements of a yeast chromosome.

2. DNA supercoiling

• Cellular DNA is highly compacted

• High degree of structural organization

• Folding mechanism– Packing DNA– Permit access to the information in the DNA

• Supercoiling

– It is an intrinsic property of DNA tertiary structure• Replication and transcription require a separation of DNA strands

• Topology: study of the properties of an object that do not change under continuous deformations– Continuous deformation: conformational changes– discontinuous deformation: involve DNA strand breakage

FIGURE 24-10 Supercoils.

FIGURE 24-11 Supercoiling of DNA.

FIGURE 24-12 Supercoiling induced by separating the strands of a helical structure.

Most cellular DNA is underwound.• Closed-circular DNA: no breaks in either strand

– B-form structure– Relaxed than supercoiled– Supercoiling results when DNA is subject to the structural strain– DNA structure 는 strain 을 받아서 supercoil 을 형성하고 그 strain 은

세포에 의해서 조절된다 .– Underwinding

• Strain is a result of an underwinding.• Fewer helical turns (84/8=10.5 bp/turn 84/7=12 bp/turn)

– Deviation from the most stable DNA form (B form)– Thermodynamically strained– Separation (10bp: access to the information) or supercoiling

(compact)– Cells actively underwind DNA with the aid of enzymatic processes

• Only when the DNA is a closed circle• When it is bound and stabilized by proteins • So that the strands are not free to rotate about each other• Nick: relaxed state

FIGURE 24-13 Relaxed and supercoiled plasmid DNAs.

FIGURE 24-14 Effects of DNA underwinding.

FIGURE 24-15 Linking number, Lk. ( 연결수 )

FIGURE 24-16 Linking number applied to closed-circular DNA molecules.

underwinding

Separation or supercoiling

= -0.01 (1%)

Strands can be unraveled and separatedNo topological bond exists

2,100 bp/10.5 = 200

FIGURE 24-17 Negative and positive supercoils.

Topoisomers: 2 forms of a given circular DNA that differ only in a topological property such as linking number No effect on the number of vase pair, atoms

FIGURE 24-18 Ribbon model for illustrating twist and writhe.

FIGURE 24-19 Promotion of cruciform structures by DNA underwinding.

FIGURE 24-20 Visualization of topoisomers.

Changes in Lk catalyzed by Type I topoisomerases: change Lk in increments of 1

Supercoiled DNA is more compact

MECHANISM FIGURE 24-21a Bacterial type I topoisomerases alter linking number.

E. coli’s topoisomerases

• I through IV• Type I (topoisomerase I and III)

– generally relax DNA by removing negative supercoils (increasing Lk)

• Type II (topoisomerase II or DNA gyrase)– introduce negative supercoils (decrease Lk)– Uses the energy of ATP– Cleave both strands of DNA

• The degree of supercoiling of bacterial DNA is maintained by regulation of the net activity of topoisomerase I and II

MECHANISM FIGURE 24-21b Bacterial type I topoisomerases alter linking number.

MECHANISM FIGURE 24-21c Bacterial type I topoisomerases alter linking number.

Eukaryotic cell’s topoisomerases

• I and II• Type I (topoisomerase I and III) : 1 가닥을 끊어서 긴장을 품

• Type II (topoisomerase II and II) : 2 가닥 모두 끊어서 긴장을 품

– Cannot underwind DNA, although both can relax both positive and negative supercoils

– Cannot introduce negative supercoils– Negative supercoils in eukaryotic cells ?

→ 24.3 (slide 54)

FIGURE 24-22 Proposed mechanism for the alteration of linking number by eukaryotic type IIA topoisomerases. 1

<Quinolone antibiotics> Bacterial type II topoisomerase, DNA gyrase inhibitor

<Camptothecin derivatives> Eukaryotic type I topoisomerase inhibitor : anti-cancer

Type II topoisomerase inhibitor: antitumor

(adriamycin)

DNA compaction requires a special form of supercoiling

• Plectonemic (twisted and thread): 꼬인 실– The supercoils are right-handed in a negatively

supercoiled DNA molecules– Extended and narrow rather than compacted– Multiple branches– General structure of supercoiled DNA in solution

• Solenoidal– Tight left-handed turns– Negative supercoiling (both)– Underwound DNA (both)– Stabilized by protein binding– Readily interconvertible– Great degree of compaction

FIGURE 24-23 Plectonemic supercoiling.

FIGURE 24-24 Plectonemic and solenoidal supercoiling.

FIGURE 24-24 Plectonemic and solenoidal supercoiling.

3. The structure of chromosome

• Chromosome ( 염색체 )– 유전정보의 저장고가 되는 nucleic acids– 색소로 진하게 염색되는 물체 ( 염색체 )– 유사분열 (mitosis) 사이 , 직전에 나타남 ( 농축됨 )

• Chromatin ( 염색질 ): chromosomal material– 정지되어 있는 분열하지 않는 세포상에서 염색체 물질

– 비정형이고 핵 중에 무질서하게 분산

– Consist of fibers containing protein and DNA, RNA (little)– DNA is tightly associated with Histones– Nucleosomes: structural unit

The beads: 8 histone molecules (2 copies of H2A, H2B, H3, H4)

Spacing: 200 bp (146 bp in histone core, remainders are linker DNA between nucleosome)

FIGURE 24-26 Nucleosomes.

Histones are small, basic proteins

• Found in chromatic of all eukaryotes• Mr. 11,000 to 21,000• Very rich in the basic amino acids arginine and lysine (25%)• 5 major classes (Mr, amino acid composition)

– H3, H4: nearly identical in aa sequences in all eukaryotes: strict conservation of their functions

– H1, H2A, H2B: less sequence similarity among species• Variants forms by methylation, ADP-ribosylation,

phosphorylation, acetylation– Affect the net electric charge, shape, other properties of

histones, as well as the structural and functional properties of chromatin

– Regulation of transcription

TABLE 24-4 Types and Properties of Histones

Nucleosomes are the fundamental organizational units of chromatin

• The compaction• Beads-on-a-string arrangement: complexes of histones

and DNA: nucleosome– The beads: 8 histone molecules (2 copies of H2A, H2B,

H3, H4)– Spacing: 200 bp (146 bp in histone core, remainders are

linker DNA between nucleosome)– Preferential degradation of the linker DNA release

histones containing 146 bp DNA– 8 histone molecules with the DNA wrapped in a left-

handed solenoidal supercoil

FIGURE 24-27a DNA wrapped around a nucleosome core.

(a)

FIGURE 24-27a DNA wrapped around a nucleosome core.

(b)

FIGURE 24-27a DNA wrapped around a nucleosome core.

(c)

FIGURE 24-28 Chromatin assembly.

Chromatin assembly

Relaxed, closed circular DNA

Binding of a histone core induces one negative supercoil and one positive supercoil

Relaxation by topoisomerase

Positioning of a nucleosome to make optimal use of A=T compression of the minor groove

Nucleosomes are packed into successively higher-order structures

• Wrapping DNA around a nucleosome core: 7-fold compaction

• Nucleosome packing– 30 nm fiber– H1 per nucleosome core required for packing– transcription region: less-ordered state, little H1– 2nd level of chromatin organization: 100-fold

compaction of the DNA• Nuclear scaffold: separated by loop of DNA with 20,000 to

100,000 bp• Additional layers of organization: coils upon coils upon

coils……………………

FIGURE 24-30 The 30 nm fiber, a higher-order organization of nucleosomes.

The 30 nm fiber, a high-order organization of nucleosomes

FIGURE 24-32 Loops of chromosomal DNA attached to a nuclear scaffold.

separated by loop of DNA with 20,000 to 100,000 bp

FIGURE 24-33 Compaction of DNA in a eukaryotic chromosome.

Active chromatin: sensitive to digestion by nuclease: hypersensitive site >>>Euchromatin

Inactive chromatin: densely packed >>>> heterochromatin: constitutive( always condensed) facultative ( at times condensed)

Condensed chromosome structures are maintained by SMC proteins

• 3rd major class of chromatin proteins• SMC proteins: structural maintenance of chromosomes• 5 distinct domains

– N, C: part of ATP hydrolytic site– 2 regions of -helical coiled-coil motifs– A hinge domain

• Dimer, V-shaped• From bacteria to human• Eukaryotes

– Cohesins: linking sister chromatids after replication – Condensins: condensation of chromosomes during

mitosis, condensin binding cause DNA overwound (positive supercoil)

FIGURE 24-34 Structure of SMC proteins.

Bacterial DNA is also highly organized

• Bacterial DNA is compacted in a structure, called the Nucleoid– Attached at one or more points to the inner surface of

the plasma membrane– Scaffoldlike structure– Organize the circular chromosome into a series of

looped domains– HU (Mr, 19,000): histonelike proteins– Relatively dynamic, more ready access to its genetic

information

FIGURE 24-36 E. coli nucleoids.

FIGURE 24-37 Looped domains of the E. coli chromosome.

FIGURE 24-35 Model for the roles of cohesins and condensins during the eukaryotic cell cycle.

Single Nucleotide Polymorphism (SNP)

• Common DNA sequence variations among individuals in genome wherein the least frequent allele has an abundance of 1% or greater.

• Make up about 90% of all human genetic variation • Occurs every 100 bases along the 3-billion-bases human

genome (dbSNP 132 version : 30,443,445 SNPs)• Some SNPs are reported to be highly related to diseases

or influence cells response to a drug

Person 1….ATCCTGTACCTACGTGTACAATAGTA…..CTGATCATCTCTATGGG….Person 2….ATCCTGTTCCTACGTGTACAATAGTA…..CTGATCATCTCTATGGG….Person 3….ATCCTGTACCTACGTGTACAATAGTA…..CTGATCAGCTCTATGGG….

SNP1 SNP2

Epigenetics and Different Aspects of Life

• Development of multicellular organism• Environment-organism interaction For examples: Nutrition supplements and environmental toxins

Image: Randy Jirtle

• Pathogenesis of diseases

SNP Types

regulatory region SNP

intron SNP Coding region SNP

Synonymous SNP

Intergenic SNP

Gene Gene

Amino acid Substitution

No amino acid substitution

Possible protein function Change

Non-synonymous SNP

Splicing region SNP

Relation between SNP and Mutation

Genome wide association study

• An examination of most of the SNPs of different individuals of a particular species to see how much the SNPs vary from individual to individual.

• Goal : Uncover the genetic basis of a given disease.• Basic Idea : A rather vague idea of a study design that

involves genotyping cases and controls at a large number of SNP markers spread throughout the genome. Look for associations between the genotypes at each locus and disease status

Case Control

Comparison of SNP Genotype frequency

Epigenetics

• Epigenetics refers to the study of changes in the regulation of gene activity and expression that are not dependent on gene DNA sequence.

• While epigenetics often refers to the study of single genes or sets of genes, epigenomics refers to more global analyses of epigenetic changes across the entire genome.

Epigenetics Mechanisms

Gene Expression

RNA Interference

Histone Modifications DNA Methylation

DNA Methylation

http://www.cellscience.com/reviews7/Taylor1.jpg

HypomethylationHypermethylation

Tissue-specific DNA methylation

Nature Genetics, 38, 1359-1360 (2006)

DNA Methylation and Cancer

Robertson, Nature Reviews Genetics, Vol6, 597

DNA Methylation and Other Human Diseases

-- Imprinting Disorder:• Beckwith-Wiedemann syndrom (BWS)• Prader-Willi syndrome (PWS)• Transient neonatal diabetes mellitus (TNDM)

-- Repeat-instability diseases• Fragile X syndrome (FRAXA)• Facioscapulohumeral muscular dystroph

-- Defects of the methylation machinery• Systemic lupus erythemtosus (SLE)• Immunodeficiency, centromeric instability and facial anomalies (ICF) syndrome

Histone Modifications

http://porpax.bio.miami.edu/~cmallery/150/gene/c7.19.4.histone.mod.jpg

Histone Modifications

http://www.nature.com/nsmb/journal/v14/n11/images/nsmb1337-F1.gif

Li e. al. (2007) Cell 128, 707

SIRT1 activator: Resveratrol analogy Resveratrol is found in grapes, wine, grape juice, and berries of Vaccinum species including blueberries, bilberries, raspberries( 覆盆子 )and cranberries. Cell energy

demand [ATP]/[AMP]

Activationde l’ AMPK Resveratrol

SIRT 1

FOXO PPARPGC-1 α

EnergeticMetabolism

Release of fattyacids from adipose

tissue

Calorie Restriction

Food Serving Total resveratrol (mg)Peanuts (raw) 1 c (146 g) .01-0.26Peanuts (boiled) 1 c (180 g) .32-1.28Peanut butter 1 c (258 g) .04-.013Red grapes 1 c (160 g) .24-1.25

(SIRT1: histone deacetylase, HDAC 의 일종 )

Deacetylation 시켜 활성유지

요약

1. Gene, central dogma, prokaryote, eukaryote 2. Intron, exon, repetitive sequence3. Centromere, telomere4. Supercoiling - topoisomer - type , topoisomerase, inhibitorsⅠ Ⅱ5. Nucleosome, Euchromatin/heterochromatin6. 30nm chromatin fiber 7. SMC protein8. SNP 9. Epigenetics/epigenome: DNA methylation, histone modification