Download Central dogma of molecular biology

Central dogma of molecular biology
•  Settled opinion
•  Principle or tenet
•  Doctrine laid
down with
authority
Central dogma of molecular biology
Central dogma of molecular biology
Central dogma of molecular biology
Nucleic acids
Central dogma of molecular biology
amino acids
Stuff responsible for hereditary
information
•  Must be stable
–  Information that makes life GO has stood
the test of time (evolution)
•  Must replicate accurately
–  Most mutations are deleterious
•  Must be capable of change
–  Without change (something of a lottery)
there would be no variability, and no
evolution
DNA (and RNA) are nucleic acids
A, G, C, T/U
•  polymers consisting of
monomers termed
nucleotides.
•  nucleotides: a molecule
composed of:
–  a pentose sugar
–  a phosphate group
–  and an organic
molecule called a
nitrogenous base.
Polynucleotide
Phosphodiester Backbones
Mb long
“Bases”
Adenine
Thymine
Guanine
Cytosine
3’!
5’!
Phosphodiester Bonds
Note 5’- 3’Orientation
Bases: DNA
34 5
2 1 6
6 5
1
2 3 4
7
8
9
Pyrimidines (one ring,
long name)
Bases: DNA
Bases: DNA
Purines (two rings, short
name)
Chargaff’s Rules
•  Base composition varies among
species
•  BUT: there are regularities.
•  The amount of guanine always
equals the amount of cytosine.
•  The amount of thymine always
equals the amount of adenine.
•  The amount of adenine +
guanine = 50% of the total.
(So 50% of the bases are
purines).
•  The amount of cytosine +
thymine = 50% of the total.
(So 50% of the bases are
pyrimidines).
Using Chargaff’s rules
•  The % of A in DNA isolated from human liver
is observed to be 30%. What is the expected
percentage of
•  (a) T
•  (b) G
•  (c) C
Using Chargaff’s rules
•  The % of A in DNA isolated from human liver
is observed to be 30%. What is the expected
percentage of
•  (a) T
Because the amount of A=T, then amount of T should
be ~30%.
•  (b) G
•  (c) C
G+C together make up the remainder:
100% - 30% -30% = 40%, divided by 2 = 20%
20%
Base pairing
The DNA molecule is a helix
•  X-ray
crystallography
revealed a helical
structure.
•  This beautiful
image was taken
and analyzed by
Rosalind Franklin.
Book alert!
Aha!
•  Watson and Crick
formed a great
intuitive partnership.
•  They depended on A
LOT of other work.
•  They could see the
forest for the trees.
DNA structure
5’ → 3’
5’ → 3’
DNA structure
5’ → 3’
5’ → 3’
DNA structure
5’ → 3’
5’ → 3’
Bases: DNA
Bases: RNA
DNA nucleotides
RNA nucleotides
RNA nucleotides
Compare DNA and RNA
•  Why 2’ OH in
RNA?
•  Why is RNA
single stranded?
•  Is RNA always
single stranded?
•  Why Uracil
instead of
Thymine in
RNA?
RNA structure
•  Inherently unstable
because the 2’-OH
group is reactive, and
so the backbone of
RNA breaks down
pretty easily
•  Except where RNA is
a genome (some
viruses) RNA
molecules are shorter
than DNA, and
transient.
RNA structure
•  RNA molecules are less
stable BUT can make some
pretty fancy secondary
structures (by internal
complementarity) that can
be catalytic.
RNA structure
•  Why the 2’-OH in RNA?
–  RNA is transient; the stored information is stable (DNA) but the
expressed information is transient (RNA) – because you don’t
need all the information all the time…
•  Is RNA always single stranded?
–  No! Sometimes it can form very elaborate secondary structures,
which can even be catalytic! In other words, you don’t always need
protein to catalyze things…maybe once upon a time, information
storage, information propagation and information expression were
all carried out by the same molecule (“RNA world”)
–  The 2’-OH increases the stability of of an intra-stand duplex by
locking it into a more compact A-from helix and may also be
important in RNA-protein interactions
•  Why Uracil instead of Thymine in RNA?
–  …later….
RNA function
DNA
RNA
mRNA
tRNA
protein
rRNA
Central dogma of molecular biology
Nucleic acids
Central dogma of molecular biology
Permanent chemical
carrier of genetic
instructions
GENOTYPE
Nucleic acids
Short-term
copy of
genetic
instructions
Genetic information is
copied over
(transcribed)
Everything is protein,
or is made by protein:
PHENOTYPE
Translation from
one chemical
language to another
DNA replication (synthesis)
•  Double stranded anti-parallel molecule.
•  Know the structure of the bases in the
context of a nucleotide (deoxyribose nucleic
acid).
•  Understand how DNA structure differs from
RNA structure, and why.
•  Learn how DNA is replicated.
•  Learn how DNA is proof-read.
•  What kinds of errors can occur?
•  What are the consequences of these errors?
How does DNA replicate?
Semi-conservative model
The DNA double helix is anti-parallel
It can be opened up to create
templates for copying
•  Single stranded
DNA can serve as a
template for highfidelity replication.
•  Also as a template
for making mRNA
(transcription).
Cell division would go nowhere
without DNA replication
Cell division would go nowhere
without DNA replication
Cell division would go nowhere
without DNA replication
One replicated chromosome
(consisting of two sister
chromatids)
One (unreplicated) chromosome
5’-AAAGGCTGATCA-3’!
’
’!
5 -AAAGGCTGATCA-3
3’-TTTCCGACTAGT-5’!
3’-TTTCCGACTAGT-5’!
S phase
5’-AAAGGCTGATCA-3’!
3’-TTTCCGACTAGT-5’!
Mitosis, meiosis I
Chromosomes are LONG pieces of
DNA
Origins of replication: multiple points
along a chromosome where replication begins
Origins of replication: multiple points
along a chromosome where replication begins
Replication has to plough through chromatin
Level 1:
nucleosome formation
Level 2:
30 nm fiber
Level 3:
Nuclear scaffolding
Level 4:
Mitotic (metaphase)
chromosome
What do you need for DNA
replication?
• 
• 
• 
• 
Template
Enzymes
Primer
dNTP’s
(d=dexyribose, N=
base A,T,G,C)
•  Energy
Proteins for DNA replication:
Single-stranded binding proteins
•  Proteins
that
stabilize the
unwound
single
stranded
DNA
Enzymes for DNA replication:
Helicases
•  DNA helicases.
•  Enzymes that unwind
the DNA double helix
for DNA replication
•  proteins bind to
specific DNA
sequences.
•  Dna/A, Dna/B, Dna/C.
Enzymes for DNA replication:
Primase
•  Primase: provides a short,
complementary strand of
RNA that is required for
DNA synthesis from a naked
DNA template.
•  There is no known DNA
polymerase that can initiate
synthesis of a DNA strand –
they can only add nucleotides
to a pre-existing strand.
•  Don’t confuse a template
with a primer.
DNA polymerase needs the 3’OH
Enzymes for DNA replication:
DNA polymerase III
•  DNA polymerase III
•  The enzyme that adds
complementary
nucleotides to the
backbone, based on
the sequence of the
single stranded
template.
•  Can only work 5’ to 3’.
Enzymes for DNA replication:
DNA polymerase III
•  Directionality of
synthesis due to
polarity of bond:
incoming dNTP needs
3’OH to react with
high energy
phosphate bond.
Enzymes for DNA replication:
DNA polymerase III
•  Directionality of
synthesis due to
polarity of bond:
incoming dNTP needs
3’OH to react with
high energy
phosphate bond.
Energy for reaction
carried on 5’
triphosphate of
incoming free
nucleotide
Enzymes for DNA replication:
DNA polymerase III
Energy for reaction
carried on 5’
triphosphate of
incoming free
nucleotide
•  Directionality of synthesis due to polarity
of bond: incoming dNTP needs 3’OH to
react with high energy phosphate bond.
Central dogma of molecular biology
nucleic acids
Transcription
•  RNA molecules
shorter than
genomes! (But
there are such
things as RNA
genomes!)
•  transient
•  NO primer
•  Proofreading?
Different types of RNA
Other kinds of RNA: dsRNA, siRNA, microRNA, pRNA etcRNA (I made this one up…)
HUGE burgeoning field…
Central dogma of molecular biology
amino acids
Protein structure – very diverse
Protein structure – why so diverse?
•  A protein is a chain of
amino acids, and there
are twenty amino acids
(that we know of…).
•  Amino acids have
unique electrostatic
and chemical
properties.
•  There are energetically
favorable ways amino
acid chains can fold…
A chain of amino acids makes a
polypeptide
A chain of amino acids makes a
polypeptide
Polypeptide chains fold into higher
order structures called proteins
•  DNA is nucleic acid.
•  RNA is nucleic acid.
•  Proteins are made up of amino acids.
•  How do you turn nucleic acids into
amino acids????
The Genetic Code: nucleic acids
spell amino acids with three letters
The Genetic Code
U’s
instead of
T’s
The Genetic Code
We’re in
triplets!
The Genetic Code
Special
spellings
for
START
and STOP
The Genetic Code
More than
one way of
spelling an
amino acid
The code is
degenerate
The Genetic Code
•  Triplet, unpunctuated and non-overlapping.
–  Reading frame depends where coding starts.
Triplets in RNA are codons.
•  Degenerate: most amino acids are specified by
more than one codon.
–  In fact only Met and Trp are specified by a single
codon.
•  Degeneracy is usually found only in the third
nucleotide of the codon.
•  The code is unambiguous: no codon specifies
more than one amino acid.
–  Exceptions: UAA, UGA and UAG.
•  The code is universal.
–  Some organelle variants, some nuclear variants.
Where does the code fit in?
DNA
RNA
mRNA
tRNA
protein
rRNA
tRNA’s match the right amino
acid with the right codon
Read about
“wobble”
Protein synthesis takes place in
the ribosome
http://www.youtube.com/watch?v=D3fOXt4MrOM&feature=related
The Ribosome is a ribozyme!!!
The Ribosome is a ribozyme!!!
The Ribosome is a ribozyme!!!
The Ribosome is a ribozyme!!!
Reading frames
Reading frames
Reading frames
The first methionine in
bacterial translation is
N-formylmethionine
Where to start?