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Chapter 1.

DNA and chromosome


Tiana Milanda

Chemical reactions of life

Metabolism
forming

bonds between molecules

dehydration synthesis = building polymers anabolic reactions

breaking

bonds between molecules

hydrolysis = digesting polymers catabolic reactions

Chemical reactions & energy

Some chemical reactions release energy


exergonic

: - digesting polymers - hydrolysis/catabolism

Some chemical reactions require input of energy


endergonic

- building polymers - dehydration synthesis/ anabolism G = change in free energy = ability to do work

Energy & life

Organisms require energy to live


where

does that energy come from?

coupling exergonic reactions (releasing energy) with endergonic reactions (needing energy)

energy

energy

Phosphate

PO4
P

bound to 4 O connects to C through an O PO4 are anions with 2 negative charges function of PO4 is to transfer energy between organic molecules (ATP)

Exergonic = spontaneous reactions?

If reactions are downhill, why dont they just happen spontaneously?


because

covalent bonds are stable


Why dont polymers (carbohydrates, proteins & fats) just spontaneously digest into their monomers

Activation energy

the amount of energy needed to destabilize the bonds of a molecule


moves

the reaction over an energy hill

Got a match? No, thats too much energy to get the work of life done!

Catalysts

So whats a cell to do to reduce activation energy? get help! ..chemical help catalyst ENZYMES

Call in the... ENZYMES!

Enzymes & substrates


substrate

reactant which binds to enzyme enzyme-substrate complex: temporary association

product

not consumed in reaction

- single enzyme molecule


can catalyze thousands or more reactions per second - enzymes unaffected by the reaction

Proteins

Structure / monomer
20

amino acids levels of structure

Function
enzymes transport

signals

defense u structure u receptors


u

peptide bond

Examples
digestive

enzymes, membrane channels, insulin hormone, actin

Genetic Code of Proteins

DNA

Evidence that DNA is the inherited genetic materials


In 1869, Friedrich Miescher (Swiss) isolated nuclein from white blood cells and found that nuclein could not be degradated by proteases nuclein was not made from proteins 2. Walter Flemming identified that materials had acidic properties, which led nuclein to be renamed nucleic acid. 3. Subsequent studies determined two major types of nucleic acids : DNA and RNA. 4. Frederick Griffiths experiment (1928)
1.

Evidence that DNA is the inherited genetic materials


Griffith was studying two strains of Streptococcus pneumoniae, a microbe that causes pneumonia
Heat-killed, diseasecausing bacteria (smooth colonies)

Disease-causing Harmless bacteria bacteria (smooth (rough colonies) colonies)

Heat-killed, diseasecausing bacteria (smooth colonies)

Control (no growth)

Harmless bacteria (rough colonies)

Dies of pneumonia

Lives

Lives Live, disease-causing bacteria (smooth colonies)

Dies of pneumonia

Evidence that DNA is the inherited genetic materials


Griffiths Hypothesis :

The genetic material from the nucleus of the dead smooth bacteria (disease causing bacteria) had transformed into the rough bacteria (harmless bacteria) to disease-causing ones. From this experiment, biologists inferred that genetic material could be transferred from one bacterium to another capsule ? The uptake of the genetic material by the cells called transformation

Evidence that DNA is the inherited genetic materials


5. Avery, MacLeod and McCartys experiment (1944)

The extract from the dead smooth bacteria was treated by protease, RNase and DNase. These treated extracts were mixed with the living rough bacteria and injected into mice. Result : DNase-treated extract could not transform rough cells to smooth cells, because DNA in these mixture was degradated by DNAse evidence that the inhereted genetic materials is DNA.

Evidence that DNA is the inherited genetic materials


6. Hershey-Chases experiment (1952)

Bacteriophage with phosphorus-32 in DNA

Phage infects bacterium

Radioactivity inside bacterium

Bacteriophage with sulfur-35 in protein coat

Phage infects bacterium

No radioactivity inside bacterium

Conclusion :

Nucleic acids is the inhereted genetic materials.

Two major types of nucleic acid :


hereditary information DNA (deoxyribonucleic acid) transmit hereditary information RNA (ribonucleic acid)
store

Building block of DNA

DNA is made up of a series of monomers called nucleotides. Each nucleotide has three parts :
a

nitrogenous base : interchangable components a pentose sugar (5C) deoxyribose in DNA (ribose in RNA) a PO4 group

Types of nitrogen bases


2 types of nitrogen bases :

Purines, double ring N base : adenine (A) guanine (G) Pyrimidines, single ring N base : cytosine (C) thymine (T) in DNA (uracil (U) in RNA)

5-Methylcytosine (5mC)

Nomenclature
Base
Purines adenine

Nucleoside (+ deoxyribose)
deoxyadenosine

Nucleotide (+ phosphate)
deoxyadenosine 5 monophosphate, dAMP, A deoxyguanosine 5 monophosphate, dGMP, G

guanine

deoxyguanosine

Pyrimidines thymine cytosine

deoxythymidine
deoxycytidine

deoxythymidine 5 monophosphate, dTMP, T deoxycytidine 5 monophosphate, dCMP, C

Polinucleotide

Backbone

the sugar of one nucleotide connect (C3) to the PO4 group (C5) of an adjacent nucleotide = phosphodiester bond
N bases hang off the sugar-phosphate backbone single nucleotide chain (C1)

The bases

Each strand of polinucleotide has polarity : 5 end (PO4 end) 3 end (OH end)

Structure of a polyii). Structure of the nucleotide chain DNA double helix primary structure of DNA
Hydrophobic

5
Hydrophilic

3
pApCpGpT = pACGT polynucleotide chain 3,5-phosphodiester bond

DNA Structure
Edwin Chargaff (1940) isolated DNA from the variety of different species. Results : 1. DNA yang diisolasi dari berbagai jaringan mempunyai jenis basa yang sama : adenin (A), sitosin (C), guanin (G) dan timin (T). 2. Komposisi basa bervariasi antar spesies 3. Pada semua DNA, A=T dan G=C atau A+G = T+C DNA untai ganda, kecuali pada virus tertentu DNA untai tunggal. Chargaffs rules.

DNA structure
Watson & Crick (1953) : each DNA molecule consists of two strands that join together and wrap around each other to form a double helix secondary structure of DNA

DNA structure
The two strands of a DNA molecule are joined by H bonds between complementary base pairs in opposite strand
purine A

:: pyrimidine

:: T 2 H bonds G :: C 3 H bonds (Chargaffs rule !!!)

Two strands of DNA are considered antiparalel, because the polarity of each strand is reversed relative to each other
Minor groove

Major groove

B structure DNA turned right

5 3

3 5

DNA structure

DNA structure

Mauric e Wilkins

X-ray diffraction showed that the DNA molecule is shaped like a helix and contains two (2) strands

Interesting note

Ratio of A-T : G-C affects stability of DNA molecule


2

H bonds vs 3 H bonds biotech procedures

more G-C = need higher T to separate strands

high T organisms many G-C

parasites many A-T (dont know why)

Denaturation/Dissociation of DNA
Double-stranded DNA Strand separation and formation of single-stranded random coils Extremes in pH or A-T rich regions high temperature denature first

Cooperative unwinding of the DNA strands (random coil)

Electron micrograph of partially melted DNA

Double-stranded, G-C rich DNA has not yet melted

A-T rich region of DNA has melted into a single-stranded bubble

A-T rich regions melt first, followed by G-C rich regions

Hyperchromicity
Absorbance maximum for single-stranded DNA Absorbance maximum for double-stranded DNA

Absorbance

220

260

300

The absorbance at 260 nm of a DNA solution increases when the double helix is melted into single strands.

DNA melting curve


Percent hyperchromicity
100

50

0 50 70 Temperature oC 90

Tm (melting temperature) is the temperature at the midpoint of the transition

DNA reassociation (renaturation)


Double-stranded DNA

Denatured, single-stranded DNA

k2
Slower, rate-limiting, second-order process of finding complementary sequences to nucleate base-pairing

Faster, zippering reaction to form long molecules of doublestranded DNA

Types of cells
1. Prokaryotic cell (prokaryotes) : no nucleus DNA (usually a single circular molecule) in nucleoid region and attached to plasma membrane, without a membrane separating it from rest of cell and in small circular DNA molecules 2. Eukaryotic cell (prokaryotes) : contain a membrane-enclosed nucleus and many organelles DNA (as chromatin) in nucleus, a membraneenclosed organelle (nuclear envelope), in mitochondria, chloroplast and small circular DNA molecules (plasmids)

Prokaryotic cells

Cell wall : rigid cell that protect the cell; ribosomes : organelle for protein synthesis; flagellum : a tail-like structure that they use for locomotion; capsule : smooth coat of proteins and sugars, etc.

Animal cells

Plant cells

DNA packing : chromosome


Bacterial DNA packing Human DNA packing Genetic variation

Bacterial DNA packing

Single circular chromosome


haploid

naked

DNA

no histone proteins

million base pairs

~4300 genes 1/1000 DNA in eukaryote

Plasmids

This will be important!

Plasmids

Plasmids
small

supplemental circles of DNA

5000 - 20,000 base pairs self-replicating

carry

extra genes

2-30 genes

can

be exchanged between bacteria

bacterial sex!! rapid evolution antibiotic resistance

can

be imported from environment

Human DNA packing


How do you fit 3 billion bp (base pair) of DNA (6 feet) in every human cell into nucleus?
DNA

coiling & folding


double helix nucleosomes chromatin fiber in cell is not dividing looped domains Chromosome formed during DNA replication only

Nucleosomes

Beads on a string
1st

8 histone molecules

level of DNA packing histone proteins

8 protein molecules with many positively charged amino acids (arginine & lysine) bind tightly to negatively charged DNA

Chromatin fiber/Nucleofilament structure

Looped domains

Chromosome structure

Duplicated chromosome consists of 2 sister chromatids


narrow

p-arm telomere q-arm

at their centromeres copies of the chromosomes contain identical DNA

Telomeres
Metaphase chromosome

Telomeres are protective caps on chromosome ends consisting of short

telomere

centromere

telomere
<1 to >12 kb (TTAGGG)many

5-8 bp tandemly repeated GC-rich DNA sequences, that prevent chromosomes from fusing and causing karyotypic rearrangements.

telomere structure

young

(TTAGGG)few

senescent

telomerase (an enzyme) is required to maintain telomere length in germline cells most differentiated somatic cells have decreased levels of telomerase and therefore their chromosomes shorten with each cell division

Human chromosomes

Most human cells have two sets (diploid) of 23 linear chromosomes each (23 maternal chromosomes and 23 maternal chromosomes) homologous pairs/homologues, which have a linear shape Chromosomes 1-22 autosomes, chromosomes 23 sex chromosomes, consisting of x and y chromosome Sex chromosome contain genes that influence sex traits and the development of reproductive organs Autosomes contain genes that affects other body features unrelated to sex

Homologous chromosomes

Paired chromosomes
both

chromosomes of a pair carry genes

control same inherited characters homologous = same information homologous chromosomes

diploid 2n

double stranded homologous chromosomes

Human chromosomes

male
female

Meiosis: production of gametes

Alternating processes, alternating stages


chromosome

number must be reduced


diploid haploid 2n n
humans: 46 23 meiosis reduces chromosome number

fertilization

restores chromosome number


haploid diploid n 2n

Mitosis vs. Meiosis

Differences across kingdoms

Not all organisms use haploid & diploid stages in same way
which

one is dominant (2n or n) differs but still alternate between haploid & diploid
have to for sexual reproduction

Karyotype analysis

Karyotype : ways to studying and comparing chromosome number and structure


Karyotype analysis : cells are spread on microscope slide, treated with chemicals to release and stain the chromosomes (ex. Giemsa stain), created a series of alternating light and dark bands in stain chromosomes Chromosome can be aligned and paired based on their staining pattern and their size can be used to identify genetic disease condition associated with abnormalities in chromosomes structure and number.

Human karyotype
male
female

Abnormal Chromosome Number

When nondisjunction occurs Pairs of homologous chromosomes do not separate normally during meiosis Gametes contain two copies or no copies of a particular chromosome
n+1
n+1 n1

Meiosis I

Nondisjunction

Meiosis II

Nondisjunction Gametes

n1

n+1

n 1

Number of chromosomes (a) Nondisjunction of homologous chromosomes in meiosis I (b) Nondisjunction of sister chromatids in meiosis II

Aneuploidy

Results from the fertilization of gametes in which nondisjunction occurred Is a condition in which offspring have an abnormal number of a particular chromosome If a zygote is trisomic, it has three copies of a particular chromosome If a zygote is monosomic, it has only one copy of a particular chromosome

Trisomic

Down syndrome is usually the result of an extra chromosome 21 (trisomy 21) Klinefelter syndrome is the result of an extra chromosome in a male, producing XXY individuals

Monosomic

Turner syndrome :
is

the result of monosomy X, producing an X0 karyotype

Alterations of Chromosome Structure

Breakage of a chromosome can lead to four types of changes in chromosome structure


Deletion
Duplication Inversion Translocation

Alterations of chromosome structure


(a) A deletion removes a chromosomal segment. A B C D E F G H Deletion A B C E F G H

(b) A duplication repeats a segment.

A B C D E

F G H

Duplication

A B C B C D E

F G H

(c) An inversion reverses a segment within a chromosome.

A B C D E

F G H

Inversion

A D C B E

F G H

(d) A translocation moves a segment from one chromosome to another, nonhomologous one. In a reciprocal translocation, the most common type, nonhomologous chromosomes exchange fragments. Nonreciprocal translocations also occur, in which a chromosome transfers a fragment without receiving a fragment in return.

A B C D E

F G H

M N O C D E

F G H

Reciprocal translocation M N O P Q
R

A B P

Disorders Caused by Structurally Altered Chromosomes

Certain cancers
Are

caused by translocations of chromosomes

Normal chromosome 9

Reciprocal translocation

Translocated chromosome 9

Philadelphia chromosome Normal chromosome 22 Translocated chromosome 22

Mitochondria & chloroplast

Mitochondria & chloroplasts are the organelles that convert energy to forms that cells can use for work
mitochondria:

from glucose to ATP chloroplasts: from sunlight to ATP & carbohydrates ATP = active energy carbohydrates = stored energy

ATP

ATP

Mitochondria & Chloroplasts

Double membranes = 2 membranes Grow & reproduce


semi-autonomous

organelles : moving, changing shape & dividing, that can reproduce by pinching in two organelles not part of endomembrane system

Own circular chromosome


directs

synthesis of proteins produced by own internal ribosomes

Dividing Mitochondria and Chroroplast chloroplast

Mitochondria

Who else divides like that?


What does this tell us about the evolution of eukaryotes?

Endosymbiosis theory

engulfed

1981 | ??

Mitochondria & chloroplasts were once free living bacteria


by ancestral eukaryote

Endosymbiont
cell

that lives within another cell (host)

as a partnership evolutionary advantage for both


one supplies energy the other supplies raw materials & protection Lynn Margulis

U of M, Amherst

Endosymbiosis theory
Evolution of eukaryotes

The inheritance of traits controlled by genes present in the chloroplasts or mitochondria


Depends

solely on the maternal parent because the zygotes cytoplasm comes from the egg

Some diseases affecting the muscular and nervous systems are caused by defects in mitochondrial genes that prevent cells from making enough ATP

The end