Download Information Content in Genetics:

10/16/13
Information Content in Genetics:
DNA, RNA and protein
mRNA translation into protein
(protein synthesis)
Francis Crick, 1958 [Crick,
F. H. C. in Symp. Soc. Exp.
Biol., The Biological
Replication of
Macromolecules, XII, 138
(1958)] & 1970 [Francis
Crick (1970) Central Dogma
of Molecular Biology. Nature
227:561-563] proposed
idea of Central Dogma that
was revised later by
inclusion of retrovirus
reverse transcription from
RNA to DNA and then back
to RNA.
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In prokaryotes DNA replication, DNA transcription into RNA and translation of
mRNA into proteins by ribosomes all occur in the cytoplasm. In eukaryotes DNA
replication, DNA transcription into RNA, and RNA processing all occur in the nucleus
and protein synthesis on ribosomes occurs in the cytoplasm. Eukaryotes have many
more levels of regulation in the expression of the genetic code than do bacteria.
https://www.mun.ca/biology/scarr/iGen3_05-09_Figure-L.jpg. jmp 13Sep2013
Eubacterial and eukaryotic
protein synthesis have the
same general pattern but
there is more complexity in
the eukaryote system.
http://origin-ars.els-cdn.com/
content/image/1-s2.0S0959440X09000700-sc1.jpg.
jmp 11Sep2013
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Prokaryote Protein Synthesis (mRNA
Translation) Initiation
Initiation Factors IF1 and IF3 assist the 30S ribosomal
subunit to align correctly with the mRNA with its 5’
Shine-Dalgarno sequence matching the 3’ anti-ShineDalgarno sequence of the 16S rRNA. IF2 brings the
formylMethionine-tRNA complex to pair with the mRNA
start codon AUG as IF3 falls off. Now the large 50S
ribosomal subunit with its 23S and 5S rRNAs can bind
the small ribosome-mRNA-fMet-tRNA complex in order
for protein synthesis to proceed.
http://www.nature.com/nature/journal/v475/n7354/
images_article/nature10126-f3.2.jpg. jmp 12Sep2013
http://bass.bio.uci.edu/~hudel/bs99a/
lecture23/hudel_27_18.jpg. jmp 11Sep2013
Prokaryote Protein
Synthesis Elongation
http://www.mun.ca/biology/
desmid/brian/BIOL2060/
BIOL2060-22/22_10.jpg. jmp
12Sep2013
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There is a unique aminoacyl-tRNA synthetase for
each amino acid-tRNA combination to ensure
information integrity. See figure to the right. http://
canmedia.mcgrawhill.ca/istudy3/books/0070741751/images/figures/
bro41751_1115L_lg.jpg. jmp 12Sep2013
http://bioserv.fiu.edu/~walterm/GenBio2004/
Chapter12_DNA_expression/figure%2012-07.jpg. jmp 12Sep2013
http://www.rcsb.org/pdb/education_discussion/
molecule_of_the_month/images/aaRS.gif. jmp 12SEp2013
The tRNA is the golden
molecule to the left.
Note the red spheres at
the bottom
representing the
anticodon and the top
red spheres where the
amino acid
phenylalanine is
attached to the 3’ CCA.
http://cdn.physorg.com/
newman/gfx/news/hires/
2009/31-researcherss.jpg.
jmp 12Sep2013
http://www.rpi.edu/dept/bcbp/molbiochem/MBWeb/
mb2/part1/images/protsyn.jpg. jmp 12Sep2013
Elongation Factor EF-G-GTP energized with GTP translocates
the newly elongated Peptidyl-tRNA from the A (aminoacyl)
site to the P (peptidyl) site as the old peptidyl tRNA leaves via
the E (exit) site. As GTP hydrolyzes to GDP, EF-G falls from
the A site and Elongation Factor Tu (EF-Tu) carrying a tRNA
loaded with its amino acid positions the tRNA-amino acid in A
site ensuring proper tRNA anticodon pairing with the mRNA
codon. The 23S rRNA interacts with EF-Tu to catalyze the
peptide bond between the new amino acid and the peptide
chain hydrolyzing GTP to GDP in EF-Tu so that EF-Tu and its
now empty tRNA leave the ribosome.
http://www.pnas.org/content/106/4/1063/
F1.large.jpg. jmp 12Sep2013
http://www.sciencemag.org/content/335/6074/1366/
F2.large.jpg. jmp 12Sep2013
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Paige S. Spencer & Jose M. Barral (2012)
Computational Structural Biotechnology Journal, http://
dx.doi.org/10.5936/csbj.20|204006 image at http://
journals.sfu.ca/rncsb/index.php/csbj/article/viewFile/
13/72/472. jmp 11Oct2013
Elongation Factor Tu (EF-Tu) carrying a tRNA loaded with its amino acid
positions the tRNA-amino acid in the A site ensuring proper tRNA anticodon
pairing with the mRNA codon. Wobble position effects as well as relative
amounts of a given tRNA-AA affect the rate at which the amino acid is positioned
in the A site to carry out synthesis of the peptide bond between this new amino
acid and the growing peptide in the P site.
Many antibiotics interfere with bacterial protein synthesis.
Besides treating infections, we use these antibiotics to study
key reactions in protein synthesis.
http://origin-ars.els-cdn.com/content/image/1-s2.0-S1074552110002048-gr1.jpg. jmp 11Sep2013
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http://www.pnas.org/content/
106/52/22239/F1.large.jpg. jmp
11Sep2013
The polypeptide is synthesized with its
amino (+ NH3) end leading the way
through the ribsome tunnel in the 50S
subunit. Once free of the tunnel, the
polypeptide begins to take its threedimensional shape forming domains.
http://en.wikipedia.org/wiki/
File:Ribosome_mRNA_translation_en.svg.
jmp 11Sep2013
http://www.nobelprize.org/educational/medicine/dna/
a/translation/pics/translation2.gif. jmp 12Sep2013
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Prokaryote Protein Synthesis Termination
Release Factor RF1 binds mRNA codons UAA and UAG while RF2 binds mRNA codons UAA
and UGA mimicking a charged tRNA. They separate the peptide chain from the tRNA in the P
site. Then RF3 which is a GTP-binding protein releases either RF1 or RF2 from the A site.
Now Ribosome Recycling Factor along with Elongation Factor EF-G (also a GTP-binding
protein) trigger release of RF3 and the tRNA in the Exit (E) site followed by the separation of
the small 30S subunit from the large 50S subunit. IF3 finishes the disassembly process.
http://origin-ars.els-cdn.com/content/image/1-s2.0S0959440X07001984-gr1.jpg. jmp 12Sep2013
Eukaryote Protein Synthesis
http://compbio.pbworks.com/f/central_dogma.jpg. jmp 10Sep2013
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Eukaryote 80S ribosome large 60S
and small 40S subunits from
Jonathan Dinman’s lab homepage
http://dinmanlab.umd.edu/
Crystal_80S_sep_2012.jpg. jmp 12Sep2013
The small 40S ribosome subunit
contains an 18S rRNA. The blue and red
colors show which proteins bind the
rRNA. The large 60S ribosome subunit
contains 25S joined to 5.8S and 5S
rRNAs with red, gold and mauve colors
with their respective proteins. An
assembled eukaryote ribosome is 80S so
40S + 60S = 80S. In prokaryotes, the
ribosome subunits 30S + 50S = 70S.
Just what you learned in elementary
math, right?
nonlinear function
The Svedberg unit is a coefficient for the
sedimentation rate of particles in
centrifuges. It is affected by particle
mass, density and shape. Think of the
rate of a feather falling and of a nickel
falling. [Not in a vacuum, Larry] Which
falls more quickly? Now if you glue the
nickel to the feather, what happens to the
rate of fall?
Eukaryote Protein
Synthesis Initiation
There are six initiation steps with 8 or more
protein factors that bind the initiator tRNAMethionine complex to the small 40S
ribosomal subunit (43S), followed by binding
this new complex to the mRNA (48S), which
involves scanning the Kozak sequence, after
which this complex is bound to the large 60S
http://www.nature.com/nrmicro/
ribosomal subunit (80S). eIF2 assists the
journal/v9/n12/images/
initiator tRNA-Met complex (methionine is not
nrmicro2655-f1.jpg. jmp
formylated) to bind to small ribosomal subunit
11Sep2013
with eIF2B to reactivate eIF2 by replacing its
GDP with GTP. The 5' 7-methyl guanosine cap
is recognized by eIF4 (eukaryote Initiation
Factor 4) and functions much like the ShineDalgarno sequence in prokaryotes to help
position mRNA on the initiator tRNA-Met-40S
ribosomal subunit for proper translation.
Scanning occurs for the Kozak sequence [CC]
RCCAUGG with R coding for A or G with A
being more efficient to initiate translation at
the start codon within the Kozak sequence.
eIF5 helps the large 60S ribosomal subunit
bind to this complex. [eIF6 has previously
prevented precocious binding to the complex
before the critical steps were completed. If a 3'
polyA tail is present on the mRNA, it binds
with the Poly(A)-binding protein and will fold
back over the initiation complex binding to the
5' 7mG cap binding proteins eIF-4G and
eIF-4E.
http://www.mol.biol.ethz.ch/groups/ban_group/
Initiation/Initiation?hires. jmp 11Sep2013
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http://www.nature.com/
scitable/content/ne0000/
ne0000/ne0000/
ne0000/14263719/
f4_jackson_nrm2838-f1.jpg.
jmp 11Sep2013
http://
micro.magnet.fsu.
edu/cells/
ribosomes/
images/
ribosomesfigure1.
jpg. jmp
11Sep2013
http://www.nature.com/nrmicro/
journal/v9/n12/images/
nrmicro2655-f1.jpg. jmp
11Sep2013
Eukaryote Protein Synthesis Elongation
The eukaryote elongation steps are very similar to
those in the prokaryotes. The EF-G-GTP complex
is EF2-GTP in eukaryotes. The protein elongation
factor 1 alpha (EF-1α) is a key GTP-binding
enzyme in protein synthesis which carries out the
same function as the prokaryote EF-Tu. [Small
groups of organisms in all eukaryote kingdoms use
elongation factor-like protein (EFL) for the same
purpose.] It brings the tRNA-AA complex to the
large ribosome subunit Aminoacyl (A) site for the
tRNA anticodon to pair with the mRNA codon.
Once pairing of complementary bases occurs, the
EF-1α GTPase domain catalyzes GTP to GDP and
EF-1α releases the tRNA-AA so that the amino acid
can now form a peptide bond with the amino acid
in the Peptidyl (P) site.]
Eukaryote Protein Synthesis Termination
The eukaryote release factor eRF1 recognizes all
three stop codons: UAA, UAG and UGA. Although
it is structurally different from prokaryote RF1 and
RF2 with one domain mimicking a tRNA with its
anticodon, it has the conserved gly-gly-gln
sequence needed to cleave the ester bond holding
the polypeptide to the last tRNA in the P site. A
second release factor eRF3 functions much like
prokaryote RF3 using GTP to help eRF1 release the
polypeptide and to cause the ribosomal subunits to
dissociate and fall away from the mRNA.
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Thomas Becker et al (2011) Nature Structural
& Molecular Biology 18:715-720 http://
www.nature.com/nsmb/journal/v18/n6/images/
nsmb.2057-F4.jpg. jmp 12Sep2013
Release factors in
Eukaryote Protein
Synthesis Termination
http://www.nature.com/embor/journal/v11/n3/images/embor201025-f1.jpg. jmp 12Sep2013
http://www.unifr.ch/biochem/
assets/images/dreyer/
research/sirna/image001.gif.
jmp 12Sep2013
silencing RNAs
(siRNA) and
micro RNAs
(miRNA) regulate
DNA transcription
and protein
synthesis
http://www.nature.com/npp/journal/v38/n1/
images/npp201287f1.jpg. jmp 12Sep2013
http://www.nature.com/srep/
2012/121218/srep00996/images/
srep00996-f7.jpg. jmp 12SEp2013
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Here they are
using the
bacterial protein
elongation factor
EF-Tu in the
eukaryotic
ribosome system
to study
similarities of
binding.
http://origin-ars.elscdn.com/content/image/1s2.0-S0959440X12001145gr1.jpg. jmp 11Sep2013
http://rna.ucsc.edu/rnacenter/images/
70s_atrna.jpg. jmp 11Sep2013
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http://employees.csbsju.edu/
hjakubowski/classes/ch331/
dna/genessimplecomplex.gif.
jmp 23Aug2013
Hierarchy of protein structure -- primary, secondary,
tertiary, and sometimes quaternary
Primary: linear order of amino acids with their
distinctive R groups.
Secondary: α helices and β pleated sheets held
together by Hydrogen bonding.
Tertiary: salt (ionic) bonds and disulfide (covalent)
bonds plus H bonds and hydrophobic interactions
(van der Waals forces) along with water partitioning
help hold the polypeptide in its 3-Dimensional
shape.
Quaternary: two or more polypeptides associate to
form a functional protein.
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http://www.icelandicsheep.com/
SheepMasterListPics/SRX534K-AI.jpg.
August 29, 2006
http://www.jpmoth.org/
Bombycidae/Bombycinae/
L1Bombyx_mori.jpg. August 29,
2006
http://
instruct1.cit.cornell.edu/
courses/biog105/pages/
demos/105/unit1/media/
fibroin.jpg. August 29, 2006
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Curly hair, “perms” and disulfide bonds
http://
www.rahmqvist.co.uk/
school_scissors.aspx.
1Sep2010.
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Enzymes (catalytic proteins) -- lysozyme,
amylase, catalase, trypsin, polymerases, etc
http://www.mol.biol.ethz.ch/groups/
glockshuber_group/Research/TMA/
image. August 29, 2006. amylase
Domains are regions of
specific function within a
given polypeptide.
Active site is where chemical
reaction is catalyzed.
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Francis Crick, 1958 [Crick,
F. H. C. in Symp. Soc. Exp.
Biol., The Biological
Replication of
Macromolecules, XII, 138
(1958)] & 1970 [Francis
Crick (1970) Central Dogma
of Molecular Biology. Nature
227:561-563] proposed
idea of Central Dogma that
was revised later by
inclusion of retrovirus
reverse transcription from
RNA to DNA and then back
to RNA.
17