Transcript Transcription & Translation: an overview Ribonucleic acid (RNA

CHAPTER 17 gene to protein
• George Beadle and Edward Tatum were able
to demonstrate the relationship between
genes and enzymes by studying mutants of a
bread mold, Neurospora. 17.1
• Beadle and Tatums hypothesis has been
restated as one gene-one polypeptide
Cracking the genetic code
• The first codon was deciphered in 1961 by
Marshall Nirenberg of the National Institute of
Health (NIH)
•  He synthesized an mRNA by linking only
uracil- bearing RNA nucleotides
• The artificial mRNA (poly U) was translated
into a polypeptide containing a string of only
one amino acid, phenylalanine.
• The genetic code is shared nearly universally
among living organisms.
 For example, the RNA codon CCG is translated
into proline in all known organisms.
Transcription initiation 17.7
Cast: members of transcription initiation
complex
a) transcription factors – proteins which
attach to upstream promoter to mediate RNA
polymerase II attachment
*first one goes to TATA box of promoter
b) RNA polymerase (II) - unwinds DNA and
adds RNA nucleotides in 5’ to 3’
ELONGATION
RNA polymerase II unwinds DNA and exposes
10-20 DNA nucleotides as it adds RNA
nucleotides to growing mRNA strand (about
60 per second)
Several RNA polymerase II often follow each
other to transcribe many mRNA strands at
once
TERMINATION
Termination signal (AAUAAA) on mRNA
(downstream) signals RNA polymerase II to
terminate transcription
At around 10 to 35 nucleotides past the
termination signal, the pre-mRNA is cut free
RNA polymerase II may go on past this!
mRNA processing
5’ Cap - Guanosine triphosphate added to 5’
end by enzymes in nucleus
a) protects against degradation
b) aids in ribosome attachment
3’ poly A tail - 50 to 250 adenine nucleotides
added to 3’ end by enzymes in nucleus
a) same functions as 5’ cap
b) also may assist with exit from nucleus
SPLICING pre-mRNA
RNA splicing – removal of introns so that only
exons remain (exons = mRNA that exits
nucleus (includes leader and trailer ))
Roberts and Sharp 1977
RNA polymerase II transcribes whole
transcription unit (DNA that is transcribed),
but many nucleotides need to be spliced to
form true mRNA from primary transcript
mRNA (pre mRNA)
Splicing
Cast:
A) small nuclear RNA (snRNA)
and other proteins form a snRNP (snurp)
small nuclear ribonucleoproteins
B) Spliceosome – several snRNP’s form this
complex- this is where coding regions of
introns are targeted for cutting. Cuts and
pastes.
Ribozymes- RNA molecules functioning as
enzymes
*all biological catalysts are proteins? No
Alternative Splicing
importance/?
tRNA anatomy
17.13
Aminoacyl tRNA synthetase 17.14
-20 types (why?)
-requires ATP to unite tRNA with proper
amino acid
W o bb l e
tRNA 3rd base
U
pairs with
I (inosine)
“ “
mRNA
A or G
U, C, A
Ribosome anatomy
rRNA + proteins
*rRNA is most abundant RNA
* two subunits compose eukaryotic ribosome
Prokaryotic ribosomes are unique
Importance?
TRANSLATION
I. Initiation 17.17
mRNA + small subunit + initiator tRNA +
large subunit (requires GTP) =
translation initiation complex
*initiation factors required too!
II. Elongation 17.18
codon recognition (2 GTP)
peptide bond formation
translocation (GTP)
E P A
E= exit site
P= middle, tRNA with polypeptide chain
= also holds initial tRNA during initiation
A= arrival site
3’
0 -amino
XXX
ACU -anticodon 5’
5’ xxxxxxxx UGAxxxxxxxxxxxxxxxxxxx-mRNA3’
III. Termination 17.19
release factor binds to stop codon
water is added to hydrolyze polypeptide
chain from tRNA in P site
Point mutations
I. Substitutions
a) silent mutation- does not alter protein
b) missense mutation – wrong amino acid
in place of correct one – usually affects
protein function
c) nonsense mutation- random formation
of premature stop codon
II. Insertions/deletions
a) frameshift mutations
ONE GENE  one polypeptide or RNA
molecule