Download Chapter 6 From DNA to Protein: How Cell Read the Genome

Chapter 6
From DNA to Protein:
How Cell Read the Genome
Genetic information directs the synthesis of protein
The central dogma of molecular biology
Genes can be expressed with different efficiencies
Untranscribed portions
Nucleotide
The chemical structure of RNA differs slightly
from that of DNA
3’
phosphodiester
bond
5’
Uracil forms a base pair with adenine
2 hydrogen bonds
RNA molecules can form intramolecular base pairs
and fold into specific structures
Base-pair with complementary sequences
Conventional
base-pair
interactions
Nonconventional
base-pair
interactions
Transcription produces an RNA complementary
to one strand of DNA
Coding strand
Sense strand
Non-Coding strand
Anti-sense strand
Template strand
Sense & antisense strand
DNA duplication by DNA polymerase
Transcription by RNA polymerase
DNA is transcribed by the enzyme RNA polymerase
ATP
CTP
UTP
GTP
Transcription can be visualized in the electron microscope
rRNAs
Gene 1
RNA polymerase
Gene 2
DNA
Ribosomal proteins
RNA polymerase vs DNA polymerase
RNA polymerase
DNA polymerase
Ribonucleotides
Deoxyribonucleotides
Without primer
With primer
1/104 error rate
1/107 error rate
Types of RNA produced in cells
messenger RNA
non-messenger RNA
ribosomal RNA
microRNA
transfer RNA
Signals in the sequence of a gene tell bacteria
RNA polymerase where to start and stop transcription
Bacterial RNA polymerase
Chain elongation
Bacterial promoters and terminators
have specific nucleotide sequences
that are recognized by RNA polymerase
Bacterial RNA polymerase
Some genes are transcribed using
one strand DNA as a template, whereas others are
transcribed using the other DNA strand
The direction of transcription is determined by the orientation of the promoter at the beginning of each gene
The three RNA polymerases in eucaryotic cells
mRNA
sRNAs
sRNAs
To begin transcription, eucaryotic RNA polymerase II
requires a set of general transcriptional factors
-25
TATA-binding protein
Dramatic local distortion in the DNA
Phosphorylate the tail
of RNA polymerase II
Allow the template strand
to be exposed by
ATP hydrolysis
Dephosphorylated form
Transcription initiation complex
TATA-binding protein (TBP) binds to TATA box sequences
and distorts the DNA
TBP (TATA-binding protein)
TATA box
DNA
Before they can be translated, mRNA molecules made
in the nucleus move out into the cytoplasm via pore
in the nuclear envelope
Pores in
nuclear envelope
TEM
Phosphorylation of RNA polymerase II allows
RNA-processing proteins to assemble on its tail
TFIIH
(1) Capping
(2) Polyadenylation
(3) Splicing
Eucaryotic mRNA molecules are modified by
capping and polyadenylation (1)
Start after 25 nucleotides
has been polymeized
(1) To increase the stability of the eucaryotic mRNA molecule
Functions:(2) To aid its export from the nucleus to the cytoplasm
(3) To identify the RNA molecule as an mRNA
Eucaryotic mRNA molecules are modified by
capping and polyadenylation (2)
CH3
2
1
3
4
5
Eucaryotic and bacterial genes are organized differently
Promoter
Intron is longer than exon
Most human genes are broken into exons and introns
3 exons
26 exons
Special nucleotide sequences signal
the beginning and the end of an intron
Branch point of the lariat
long
intron-exon
boundary (border)
short
R: A or G
Y: C or U
N: A or C or G or U
RNA splicing might occurred before or after polyadenylation
RNA splicing
An introns forms a branched structure during splicing
5’
Branch point of the lariat
3’
5’
Spliceosome
Small nuclear RNAs (snRNAs) + Proteins
= Small nuclear ribonucleoprotein particles
(snRNPs)
The -tropomyosin gene can be spliced in different ways
(1) Many different protein to be produced from the same gene by alternative splicing
(2) 60% of human genes probably undergo alternative splicing
A specialized set of RNA-binding proteins signal that
a mature mRNA is ready for export to the cytoplasm
Recognizes and exports
only completed mRNAs
Exon junction complex (EJC)
Mark completed RNA splices
Life times depends on
(1) Nucleotide sequence (3’ untranslated sequence)
(2) The type of cell
3’ untranslated region
Procaryote and eucaryotes handle
their RNA transcripts differently
Transcription in procaryotic or eucaryotic cells (1)
Transcription in procaryotic or eucaryotic cells (2)
Bacteria
Eucaryotic cells
RNA polymerase
Single type
Three types (I, II, III)
Accessory proteins
X
General transcription factors
Nontranscribed DNA between genes Short
Long
Small RNAs
(1) siRNA (small interfering RNA)
(2) miRNA (microRNA)
(3) piRNA (piwi-interacting RNA)
miRNA
siRNA
Amino Three-nucleotide
codons
acids
The nucleotide sequence of an mRNA is translated into
the amino acid sequence of a protein
via the genetic code
Start
codon
Stop
codons
UUU codes for phenylalanine
Marshall Nirenberg
Heinrich Matthaei
Messages of mixed repeating sequences further
narrowed the coding possibilities
Gobind Khorana
The Nobel Prize in Physiology or Medicine 1968
Interpretation of the genetic code and its function in protein synthesis
An RNA molecule can be translated
in three possible reading frames
Reading frame
Frame shift
tRNA molecules are molecular adaptors,
linking amino acids to codons
L-shape molecule
dihydrouridine
pseudouridine
Wobble base-pairing
61 codons
31 anticodons
20 amino acids
The genetic code is translated by means of two adaptors
that act one after another
Charging
1
Aminoacyl-tRNA synthase
2
Charged tRNA
Anticodon
Ribosomes are found in the cytoplasm of a eucaryotic cell
Attached to the ER
Ribosome
Free in the
cytosol
TEM
A ribosome is a large complex of four RNAs and
more than 80 proteins
(rRNA)
(rRNA)
Catalyzes the formation
of polypeptide chain
Matches the tRNA to
the codon of the mRNA
1/3
2/3
Each ribosome has a binding site for mRNA
and three binding sites for tRNA
Large subunit
peptidyl-tRNA
Large subunit
Small subunit
Exit
aminoacyl-tRNA
Small subunit
Translation takes place in a four-step cycle
Catalyzed by an enzymatic site
in the large subunit
Large subunit
Ribosomal RNAs give the ribosome its overall shape
Rate of sedimentation in an ultracentrifuge
Protein
Catalytic site for peptide bond formation
Large subunit of a bacterial ribosome
Initiation of protein synthesis in eucaryotes requires
initiation factors and a special initiator tRNA
or formylmethionine in bacteria
only this charged RNA
can binds to the P-site
E
E
P
P A
A
Start codon
E
P
A
In the final phase of protein synthesis, the binding of
release factor to an A-site bearing a stop codon
terminates translation
Stop codons
UAG
UGA
UAA
A single procaryotic mRNA molecule can encode
several different proteins
Operons
Polycistronic
Procaryote
Proteins are translated by polyribosomes
Polyribosomes (Polysomes)
Inhibitors of procaryotic protein synthesis
are used as antibiotics
The proteosome degrades short-lived and
misfolded proteins
Active site of
the proteases
Ubiquination for protein degradation
Ubiquitin
Protein production in a eucaryotic cell
requires many steps
Many proteins require additional modification
to become fully functional
Glycosylation (> 100 kinds)
An RNA world may have existed
before modern cells arose
An RNA molecule can in principle guide
the formation of an exact copy of itself
Ribozyme
Ribozyme – RNA molecules that possess catalytic activity
A ribozyme is an RNA molecule that possesses
catalytic activity
Biochemical reactions that can be catalyzed by ribozymes
Could an RNA molecule catalyze its own synthesis?
RNA may have preceded DNA and proteins in evolution