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How Cells Read The Genome: From DNA To Protein
How Cells Read The Genome: From DNA To Protein
Eukaryotes
• Promoter –(asymmetrical sequence)
• Basic core promoter –TATA box (TATAAA(A)); within 50bp
upstream of start site; found in unicellular eukaryotes
• Core promoter PLUS
•Downstream promoters
•Proximal promoter elements
• Initiator sequences
• Insulators
Gene Regulatory Networks – control the number and type of
transcripts made by a cell.
video
Simple core
promoter
Holoenzyme or
Initiation Complex
Transcription Factors Have Common DNA Binding Motifs
• Zinc finger
• Helix-turn-helix
• Leucine zipper
RNA Polymerase
Recognizes and binds to TATA box; TBP +
10 TBP associated factors; position set
TBP bends DNA ~80o and
forces open the minor
groove.
Recognizes and binds to TATA box; TBP +
10 TBP associated factors
Sequential
TBP
} TFIID or TBP
IIA
Binding IIB
Model for
IIF
Eukaryotic RNA
polymerase II
Pol IIa
preinitiation helicase
IIH
protein kinase
IIE IIF
Polymerization of 1st few NTPs and IIB IIA Pol IIa
IIH
phosphorylation of CTD leads to TATA Inr
promoter clearance. TFIIB, TFIIE and initiation complex, DNA melted at Inr
TFIIH dissociate, PolII+IIF elongates,
and TFIID + TFIIA stays at TATA. Activated PIC
Transcription initiation in the cell often requires the local recruitment of
chromatin-modifying enzymes, including chromatin remodeling complexes and
histone acetylases - greater accessibility to the DNA present in chromatin
RNA polymerase is also assisted by DNA supercoiling
Phosphorylation of the carboxy
terminal domain (CTD) of one of the
subunits of RNA PolII;
RNA polymerase II dissociates from
the transcription factors and other
protein complexes that were required
for assembly and elongation begins
• Capping
• Splicing
• Polyadenylation
RNA Capping enzymes:
• Phosphatase
• Guanyl transferase – GMP in 5’ to5’
linkage
• methyltransferase
R=A or G,Y=C or U
The Spliceosome Forms
snRNAs (U1, U2, U4, U5 and U6) and associated proteins = snRNPs
• U1 binds to the GU sequence at the 5' splice site, along with accessory
proteins/enzymes,
• U5/U4/U6 trimer binds, and the U5 binds exons at the 5' site, with U6
binding to U2;
• U2/U5/U6 remain bound to the lariat, and the 3' site is cleaved and
exons are ligated using ATP hydrolysis. The spliced RNA is released and
the lariat debranches.
Rearrangements that occur during splicing
• U1 replaced by U6
• BBP (branch binding protein) and U2
• U5 complex branch forming enzymes in
U6 and U2
?
A different aminoacyl-tRNA synthetase enzyme for each amino acid
Large subunit does???? Small subunit does ?????
Translation Initiation
This is the only tRNA that can bind to the small
ribosomal subunit by itself
Protein made in 5’ to 3’ direction,
with N-terminal end made first
General Mechanism
Proteolytic core
Cap hydrolyses
ATP; regulates exit
control
Add inhibitor
edeine
cycloheximide
0 2 4 6 8
Time (min)