1,DNA structure
2,Chromosomes
3,DNA replication in bacteria
4,DNA replication in eukaryotes
F DNA structure and replication
? Bases碱基
? Nucleosides核苷
? Nucleotides核苷酸
? 3’,5’- phosphodiester bonds磷酸二酯键
? DNA sequence
? DNA double helix
F1 DNA structure
Ba se
Su g ar
A cid
Monophosphate
Diphosphate
Triphosphate
Adenine
Guanine
Thymine
Cytosine
Uracil
Nucleoside (Adenosine)
Nucleotide (Adenosine monophosphate,AMP)
Purine
Pyrimidine
Ribose,
Deoxyribose
1’
2’ 3’
4’
5’
Pentose
1
2
3
4
5
6
Uracil
N H
N
H
O
O
U
N
N
H
N H 2
O
Cytosine
C
N H
N
H
O
O
Thymine
T
Pyrimidines嘧啶
1
2
3 4
5 6
9
7
8
N
N
N
H
N
N H 2
?Adenine腺嘌呤
A
N H
NNH
N
O
N H 2
?Guanine鸟嘌呤
G
Purines嘌呤
Bases
Ribose and deoxyribose
Nucleosides核苷
1’
2’ 3’
4’
5’
(OH)
1’
2’ 3’
4’
5’
(OH)
C1, N1 C1, N9
假尿苷（ ψ） C5:C1
5
OH
Adenosine Guanosine Cytidine Uridine
Nucleosides
胞嘧啶核苷 尿嘧啶核苷鸟嘌呤核苷腺嘌呤核苷
N
N
O H
H O
N
N
N H
2
H O
N
N
O H
H
2
N
N
N
N
N
N
N
N H
2
O
H
H
O H
H
O H
H
H O C H
2
H O C H
2
O
H
H
O H
H
O H
H
O
H
H
O H
H
O H
H
H O C H
2
O
H
H
O H
H
O H
H
H O C H
2
Nucleotides
H
A nucleotide is a phosphate ester of a
nucleoside,
dGMP
O
-
P
O
O
-
N
N
N
N
N H 2
O
H
H
O H
H
O H
H
O C H 2O
-
P
O
O
-
O
-
P
O
O
-
三磷酸腺苷 ( A T P )
AMP
ADP
ATP
Nucleotides
? cAMP ? cGMP
cAMP and cGMP
7-Methylguanylate
Covenlent structure
? In a DNA structure,the different nucleotides are
covalently joined to form a long polymer chain
covalent bonding between the phosphates and
sugars,
3,5 Phosphodiester bonds
5’
5’
3’
3’
Polymer
DNA sequence
? The DNA sequence is the sequence of A,C,G
and T along the DNA molecule which carries the
genetic information,
? 5’-ACTTTCAGACC-3’
? In a DNA strand n nucleotides long,the number
of different possible sequences(or messages) is
4n
P
R
B
3’
2’
5’
1’
A T C G A T C G
P OH
5’ 3’
5’ pApTpCpGpApTpCpG-OH 3’
5’ pATCGATCG-OH 3’
ATCGATCG
DNA sequence
DNA primary structure
DNA chain polarity:5’?3’
DNA second structure
? 1953,Watson and Crick:DNA double helix
1,2 strands are organized in an antiparallel
2,The bases on the inside and the sugar-phosphate backbones on the
outside of the double helix
3,Major groove and minor groove
4,the bases of the 2 strands form hydrogen bonds
5,Complementary base pairing:A=T,G=C
6,D=2nm,n=10,0.34nm,36o
JD Watson
F Crick
1953 Cambridge
1962
Watson and Crick
DNA double helix
Double helix
牛頓 (2000) no,201 p,36
A T
C G
Antiparally chain
DNA double helix
2.0 nm
小
沟
大
沟
Hydrogen bonds
DNA double helix
Large groove
Small groove
1 Twist = 10.5 bp
1
2
3
4
5
6
7
8
9
10
34 ?
S
ECO
NDA
RY
ST
RUCT
URE
3’
5’
5’
3’
Juang RH (2004) BCbasics
? Prokaryotic chromosomes染色体
? Eukaryotic chromosomes
? Ncleosomes核小体
? 30 nm fiber
? Radial loops放射环
F2 Chromosomes
Prokaryotic chromosomes
? The DNA in a bacterium is a supercoiled double-stranded
circular molecule that is packaged in the nucleoid region
of the cell,
? The DNA is negatively supercoled,complexed to several
histone组蛋白 -like proteins (mainly proteins HU,HSP-1
and H-NS) and organized into about 50 domain bound to
a protein scaffold支架,
? p173
Eukaryotic chromosomes
? Eukaryotic cells contain much more DNA than
prokaryotes,
? In the nucleus,the DNA is packaged into
chromosomes that consist mainly of DNA and
proteins called histones although other
nonhistone proteins (NHP) are also present,
Eukaryotic chromosomes
? Each chromosome contains a single linear
double-stranded DNA molecule,
? The length of the packaged DNA molecule
varies,
? The extensive packaging of DNA in
chromosomes results from three levels of folding
involving nucleosomes,30nm filaments and
radial loops,
Nucleosomes核小体
? The chromosomal DNA is complexed with five
types of histone组蛋白 (H1,H2A,H2B,H3 and
H4),
? These are very basic proteins,rich in Arginine
and Lysine,
? The amino acid sequences of histones are
highly conserved in evolution,
Histone octamer八聚体
? The DNA is wound round a histone octamer (two
molecules each of H2A,H2B,H3 and H4 ) to
form a nucleosome,
? The DNA between neighboring nucleosomes
(linker DNA ) binds histone H1,
? The packing ratio of nucleosomes is about 7
Ncleosome
Nucleosomes
30 nm fiber
30 nm fiber
Radial loops
? DNA polymerases
? Replication forks
? Okazaki fragments
? RNA primer
? Accessory proteins
F3 DNA replication in bacteria
Semi-conservative replication
The DNA synthesis system
1,Precursors:4 dNTPs(N=A,T,G,C) Mg2+
2,Enzyme
3,Template,dsDNA or sDNA
4,Primer,a short RNA primer
5,DNA synthesis occurs in 5’— 3’direction
E.coli DNA polymerases
1,DNA polymerases I
2,DNA polymerases II
3,DNA polymerases III
DNA polymerases common structure
DNA polymerases I
5‘-3’ exonuclease外切酶活性
3‘-5’ exonuclease,
------proofreading function校对
DNA polymerases
E.coli PolIII holoenzyme structure
?-subunit bind with template
?亚基形成一个围绕 DNA的环状钳
PolIII ? subunit X-ray structure
PolIII? subunit X-ray structure
Co m p ar is on of DN A P oly m e r as e of E.c oli
D N A pol ym erase
I II II I
St ruct ural gene pol A pol B pol C ( dn a E)
Subuni t s (num ber of di f fer ent t y pes) 1 ≧ 4 ≧ 10
M
r
103,000 8 8,000 830,000
3' → 5 ' Exonucl ease ( proof readi ng) Y es Y es Y es
5' → 3 ' Exonucl ease Y es N o N o
Pol y m e ri zat i on r at e ( nucl eot i des/ sec) 1 6- 20 40 250- 1,000
P ro cess i v i t y ( nucl eot i des added bef ore 3- 200 1,500 ≧ 50 0,00 0
pol y m eras e di ss oci at es)
Replication
DNA ligase
Replication forks
? Replication starts at a single origin,is bi-
directional
and semi-conservative,
? Each replication bubble consists of two
replication forks
E.coli DNA replication model
Replication of the bacterial
circular chromosom
Okazaki fragments
? DNA synthesis proceeds in a 5'-3' direction on each strand
of parental DNA,
? On the strand with 3'-5' orientation (the leading strand) the
new DNA is synthesized continuously,
? On the strand that has 5'-3' orientation (the lagging strand )
the DNA is synthesized discontinuously as a series of short
Okazaki fragments that are then joined together,
Okazaki fragments
RNA primer
? RNA primer is synthesized by an RNA ploymerase called
primase,
? Each Okazaki fragments requires an RNA primer(about 5
nt).DNA replication is extended by DNA polymerase III,
? DNA polymerase I (5‘-3’ exonuclease)
degrades the primers and replaces it with DNA,
? DNA ligase joins the ends of the DNA fragments togeher,
Accessory proteins
1,A helicase unwinds the double helix and single-
stranded DNA-binding protein stabilizes the single
stranded regions during replication,
2,NDA topoisomerase I 拓扑异构酶 is needed to allow the
helix to unwind without causing extensive rotation of
the chromosome,
3,DNA topoisomerase II separates the two daughter DNA
circles following replication,
Topoisomerase I
Gyrase Topo II
SSB
All the proteins
1,Cell cycle
2,Multiple replicons
3,Five DNA polymerases
4,Leading and lagging strands
5,Telomere replication
6,Replication of chromatin
F4 DNA replication in eukaryotes
Cell cycle
? In eukaryotes,the cell cycle consists of
G1,S,G2,and M phases,
? Most differences in the cycle times of different
cells are due to differences in the length of the
G1 phase,
Multiple replicons
? DNA replication occurs only in the S phase,
? It occurs at many chromosomal origins,is bi-
directional and semi-conservative,
? Sets of 20-80 replicons(The DNA replication under
the control of a single origin is called a replicon) act
as replication units that are activated in sequence,
Five DNA polymerases
? DNA polymerases α and δ replicate
chromosomal DNA
? DNA polymerase β and ε repair
DNA
? DNA polymerase γ replicates
mitochondrial 线粒体 DNA
Leading and lagging strands
? DNA polymerases α synthesizes the lagging
strand,via Okazaki fragments,
? DNA polymerases δ synthesizes the leading
strand,
? The RNA primers are synthesized by DNA
polymerases α which carries a primase subunt,
Telomere 端粒 replication
? Telomerase端粒酶,a DNA polymerase that
contains an integral RNA that acts as its
own primer,is used to replicate DNA at the
ends of chromosomes,
Telomere replication