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From DNA to Protein
Chapter 14
Hsueh-Fen Juan
Oct 23, 2012
14.1 DNA, RNA, and Gene Expression
 What is genetic information and how does a cell
use it?
 Ricin:蓖麻毒素，相當可怕又易取得，毒性來自於
它可使核醣體失去功能
The Nature of Genetic Information
 Each strand of DNA consists of a chain of four
kinds of nucleotides: A, T, G and C
 The sequence of the four bases in the strand is
the genetic information
Converting a Gene to an RNA
 Transcription
• Enzymes use the nucleotide sequence of a gene
to synthesize a complementary strand of RNA
 DNA is transcribed to RNA
• Most RNA is single stranded
• RNA uses uracil in place of thymine
• RNA uses ribose in place of deoxyribose
Ribonucleotides and Nucleotides
Ribonucleotides and Nucleotides
adenine A
HC
NH 2
N CC N
N C N CH
guanine G
O
N CC
NH
HC
NC C
N NH
2
DNA
deoxyribonucleic acid
RNA
ribonucleic acid
nucleotide
base
sugar–
phosphate
backbone
cytosine C
NH 2
HC C N
HC
thymine T
N
N CC
N
HC
N C
CH
N
guanine G
O
N CC
NH
HC
NC C
N NH 2
NH 2
HC C N
HC N C O
base pair
CH 3 C C NH
HC N C O
Nucleotide
bases of DNA
NH 2
cytosine C
C O
O
adenine A
uracil U
O
HC C NH
HC N C O
DNA has one function: It
permanently stores a cell’s
genetic information, which
is passed to offspring.
RNAs have various
functions. Some serve
as disposable copies of
DNA’s genetic message;
others are catalytic.
Nucleotide
bases of RNA
Fig. 14-3, p. 217
RNA in Protein Synthesis
 Messenger RNA (mRNA)
• Contains information transcribed from DNA
 Ribosomal RNA (rRNA)
• Main component of ribosomes, where polypeptide
chains are built
 Transfer RNA (tRNA)
• Delivers amino acids to ribosomes
microRNAs
• Endogenous approximately 23 nt RNAs
Bartel DP Cell 2009.
• Control gene expression at the post-transcriptional
level by degrading or repressing target messenger
RNAs (mRNAs)
Chang YM & Juan HF et al PNAS 2008.
Bartel DP Cell 2004.
• Involved in diverse physiological and pathological
processes.
Suzuki HI et al Nature 2009.
• Oncomirs: miRNAs which is closely related to tumor
Esquela-Kerscher A & Slack FJ. Nat Rev Cancer 2006.
9
Dicer
Drosha
Pre-miRNA
unwind
mature
miRNA
Pri-miRNA
RISC
3’UTR
5’UTR
miRNA gene
AAAAA
Translational Repression
5’UTR
3’UTR
mRNA Degradation
AAAAA
Converting mRNA to Protein
 Translation
• The information carried by mRNA is decoded
into a sequence of amino acids, resulting in a
polypeptide chain that folds into a protein
 mRNA is translated to protein
• rRNA and tRNA translate the sequence of base
triplets in mRNA into a sequence of amino acids
Gene Expression
 A cell’s DNA sequence (genes) contains all the
information needed to make the molecules of life
 Gene expression
• A multistep process including transcription and
translation, by which genetic information
encoded by a gene is converted into a structural
or functional part of a cell or body
14.2 Transcription: DNA to RNA
 RNA polymerase assembles RNA by linking
RNA nucleotides into a chain, in the order
dictated by the base sequence of a gene
 A new RNA strand is complementary in
sequence to the DNA strand from which it was
transcribed
DNA Replication and Transcription
 DNA replication and transcription both
synthesize new molecules by base-pairing
 In transcription, a strand of mRNA is assembled
on a DNA template using RNA nucleotides
• Uracil (U) nucleotides pair with A nucleotides
• RNA polymerase adds nucleotides to the
transcript
• 同DNA複製，能量來自水解高能磷酸鍵
Base-Pairing in
DNA Synthesis and Transcription
The Process of Transcription
 RNA polymerase and regulatory proteins
attach to a promoter (a specific binding site in
DNA close to the start of a gene) 這句很重要，
說明了轉錄的起始準備所有所需物質
 RNA polymerase moves over the gene in a 5‘ to
3’ direction, unwinds the DNA helix, reads the
base sequence, and joins free RNA nucleotides
into a complementary strand of mRNA (RNA聚
合酶超強多功能，不需解旋酶自己就可解)
Transcription
gene region
newly forming
RNA transcript
RNA polymerase, the
enzyme that catalyzes
transcription
DNA template
winding up
DNA template
unwinding
A RNA polymerase binds to a promoter
in the DNA, along with regulatory
proteins. The binding positions the
polymerase near a gene in the DNA.
B The polymerase begins to move along the
DNA and unwind it. As it does, it links RNA
nucleotides into a strand of RNA in the order
specified by the base sequence of the DNA.
In most cases, the nucleotide
sequence of the gene occurs on only
one of the two strands of DNA. Only
the complementary strand will be
translated into RNA.
The DNA double helix winds up again after the
polymerase passes. The structure of the
“opened” DNA molecule at the transcription site
is called a transcription bubble, after its
appearance.
Fig. 14-5a, p. 218
transcription site
5’
3’
growing RNA transcript
C What happened in the gene region? RNA polymerase
catalyzed the covalent bonding of many nucleotides to one
another to form an RNA strand. The base sequence of the
new RNA strand is complementary to the base sequence of
its DNA template—a copy of the gene. (接的方式同DNA複製)
Fig. 14-5b, p. 219
Animation: Gene transcription details
Transcription
 Many RNA polymerases can transcribe a gene
at the same time
14.3 RNA and the Genetic Code
 Base triplets in an mRNA are words in a proteinbuilding message
 Two other classes of RNA (rRNA and tRNA)
translate those words into a polypeptide chain
Post-Transcriptional Modifications
 In eukaryotes, RNA is modified before it leaves
the nucleus as a mature mRNA (真核在核內修飾)
 Introns
• Nucleotide sequences that are removed from a
new RNA
 Exons
• Sequences that stay in the RNA
Alternative Splicing
 Alternative splicing (選擇性/可變 剪接)
• Allows one gene to encode different proteins
• Some exons are removed from RNA and others
are spliced together in various combinations
 After splicing, transcripts are finished with a
modified guanine “cap” at the 5‘ end and a polyA tail at the 3’ end (cap用來幫助mRNA附著核醣
體，tail用來防水解)
Post-Transcriptional Modifications
Animation: Pre-mRNA transcript
processing
mRNA – The Messenger
 mRNA carries protein-building information to
ribosomes and tRNA for translation
 Codon
• A sequence of three mRNA nucleotides that
codes for a specific amino acid
• The order of codons in mRNA determines the
order of amino acids in a polypeptide chain
Genetic Information
 From DNA to mRNA to amino acid sequence
Genetic Code
 Genetic code
• Consists of 64 mRNA codons (triplets)
• Some amino acids can be coded by more than
one codon
 Some codons signal the start or end of a gene
• AUG (methionine) is a start codon (同時也決定
met)
• UAA, UAG, and UGA are stop codons (不決定胺
基酸)
Codons of the Genetic Code
rRNA and tRNA – The Translators
 tRNAs deliver amino acids to ribosomes
• tRNA has an anticodon complementary to an
mRNA codon, and a binding site for the amino
acid specified by that codon
 Ribosomes, which link amino acids into
polypeptide chains, consist of two subunits of
rRNA and proteins (負責催化合成長肽鍊的是核
醣體的rRNA，而非蛋白質)
Ribosomes
tRNA
14.4 Translation: RNA to Protein
 Translation converts genetic information carried
by an mRNA into a new polypeptide chain
 The order of the codons in the mRNA
determines the order of the amino acids in the
polypeptide chain
Translation
 Translation occurs in the cytoplasm of cells
 Translation occurs in three stages
• Initiation
• Elongation
• Termination
Initiation
 An initiation complex is formed
• A small ribosomal subunit binds to mRNA (小次
單元附著mRNA)
• The anticodon of initiator tRNA base-pairs with
the start codon (AUG) of mRNA
• A large ribosomal subunit joins the small
ribosomal subunit
Elongation
 The ribosome assembles a polypeptide chain as
it moves along the mRNA
• Initiator tRNA carries methionine, the first amino
acid of the chain
• The ribosome joins each amino acid to the
polypeptide chain with a peptide bond
Termination
 一條mRNA上可以有多個核醣體同時進行轉譯，此
多核醣體+mRNA之複合物稱為polysome
 轉譯能量來自mRNA之GTP-cap的高能磷酸鍵
 ATP用來將胺基酸附著在free tRNA上
 When the ribosome encounters a stop codon,
polypeptide synthesis ends
• Release factors bind to the ribosome
• Enzymes detach the mRNA and polypeptide
chain from the ribosome
Translation in Eukaryotes
Initiation
A A mature mRNA
mRNA
leaves the nucleus and
enters cytoplasm, which
has many free amino
acids, tRNAs, and
initiator small
ribosomal subunits.
tRNA
ribosomal
An initiator tRNA binds
subunit
to a small ribosomal
subunit and the mRNA.
large
ribosomal
subunit
B A large ribosomal
subunit joins, and
the cluster is now
called an initiation
complex.
Stepped Art
Fig. 14-12 (a-b), p. 222
Elongation
C An initiator tRNA
carries the amino acid
methionine, so the first
amino acid of the new
polypeptide chain will be
methionine. A second
tRNA binds the second
codon of the mRNA (here,
that codon is GUG, so the
tRNA that binds carries
the amino acid valine).
A peptide bond
forms between
the first two
amino acids
(here, methionine
and valine).
Fig. 14-12c, p. 223
D The first tRNA is
released and the
ribosome moves to the
next codon in the mRNA.
A third tRNA binds to the
third codon of the mRNA
(here, that codon is UUA,
so the tRNA carries the
amino acid leucine).
A peptide bond
forms between the
second and third
amino acids
(here, valine
and leucine).
Fig. 14-12d, p. 223
E The second tRNA
is released and the
ribosome moves to the
next codon. A fourth
tRNA binds the fourth
mRNA codon (here, that
codon is GGG, so the
tRNA carries the amino
acid glycine).
A peptide bond
forms between the
third and fourth
amino acids (here,
leucine and
glycine).
Fig. 14-12e, p. 223
Termination
F Steps d and e are repeated over and
over until the ribosome encounters a stop
codon in the mRNA. The mRNA transcript
and the new polypeptide chain are
released from the ribosome. The two
ribosomal subunits separate from each
other. Translation is now complete. Either
the chain will join the pool of proteins in
the cytoplasm or it will enter rough ER of
the endomembrane system (Section 4.9).
Fig. 14-12f, p. 223
Animation: Translation
14.5 Mutated Genes
and Their Protein Products
 If the nucleotide sequence of a gene changes, it
may result in an altered gene product, with
harmful effects
 因為密碼子有退化性，不同的密碼子也可能指向
相同胺基酸，這稱為細胞的margin of safety
 Mutations
• Small-scale changes in the nucleotide sequence
of a cell’s DNA that alter the genetic code
Common Mutations
 Base-pair-substitution (點突變)
• May result in a premature stop codon or a
different amino acid in a protein product
• Example: sickle-cell anemia
 Deletion or insertion
• Can cause the reading frame of mRNA codons to
shift, changing the genetic message
• Example: Huntington’s disease
Common Mutations
part of DNA
mRNA
transcribed
from DNA
resulting amino THREONINE PROLINE
acid sequence
A Part of the DNA, mRNA,
and amino acid sequence of
the beta chain of a normal
hemoglobin molecule.
GLUTAMATE GLUTAMATE
LYSINE
base
substitution
in DNA
altered mRNA
altered amino
THREONINE
acid sequence
PROLINE
VALINE
GLUTAMATE
deletion in
DNA
altered mRNA
altered amino
THREONINE
acid sequence
PROLINE
GLYCINE
ARGININE
LYSINE
B A base-pair substitution in
DNA replaces a thymine
with an adenine.
When the altered mRNA is
translated, valine replaces
glutamate as the sixth amino
acid of the new polypeptide
chain. Hemoglobin with this
chain is HbS—sickle
hemoglobin (Section 3.6).
C Deletion of the
same thymine causes a
frameshift.
The reading frame for the
rest of the mRNA shifts, and
a different protein product
forms. This mutation results
in a defective hemoglobin
molecule. The outcome is
thalassemia, a type of
anemia.
Stepped Art
Fig. 14-13, p. 224
Animation: Base-pair substitution
Animation: Frameshift mutation
What Causes Mutations?
 Transposable elements (轉置子)
• Segments of DNA that can insert themselves
anywhere in a chromosomes
• 幾乎所有物種都有它的存在，人類有45%DNA與其
相關
• Barbara McClintick 的各種奇怪玉米
 Spontaneous mutations
• Uncorrected errors in DNA replication
• 這很正常，DNA複製速度超快，偶爾出錯也不意外
 Harmful environmental agents
• Ionizing radiation, UV radiation, chemicals
• Thymine dimer 胸腺嘧啶二聚體
McClintock’s Transposable Elements
Mutations Caused by Radiation
 Ionizing radiation damages chromosomes,
nonionizing (UV) radiation forms thymine
dimers (胸腺嘧啶二聚體)
Inherited Mutations
 Mutations in somatic cells of sexually
reproducing species are not inherited
 Mutations in a germ cell or gamete may be
inherited, with evolutionary consequences
Transcription Assembly of RNA on unwound
Summary:
Protein
Synthesis
in
Eukaryotic
Cells
regions of DNA molecule
mRNA
processing
mRNA
rRNA
proteins
mature mRNA ribosomal
transcripts
subunits
Translation
At an intact
ribosome,
synthesis of a
polypeptide
chain at the
binding sites
for mRNA and
tRNAs
Convergence
of RNAs
tRNA
mature
tRNA
cytoplasmic
pools of
amino acids,
ribosomal
subunits, and
tRNAs
Protein
Fig. 14-16, p. 226
Animation: Protein synthesis summary