Molekulární biologie(KBC/MBIOG)
Ivo FrébortAlberts et al. (2008) Molecular Biology of the Cell, 5th ed.
Garland Science, New York
4. DNA Transcription and Translation
Central dogma of molecular biology
Genes can be expressed with different efficiencies
Chemical structure of RNA
Structure of bacterial RNA polymerase
The transcription cycle of bacterial RNA polymerase
Transcription by RNA polymerase
Promoter and terminator in prokaryotes
Direction of transcription is given by the orientation of promoter sequence
(RNA synthesis always from 5‘ to 3‘)
RNA Pol IIBlue – MgRed - Zn
Initiation of transcription in eucaryotes
Transcription factors
Transcription complex in eucaryotes
From gene to protein in eucaryotes and procaryotes
Procaryotic and eucaryotic mRNA structures
Pre-mRNA processing in eucaryotes
Introns and exons
Alternative splicing
The spliceosome reaction
The spliceosome reaction
Three types of RNA splicing
Rare self splicing intrones – evolutionary basis for spliceosomes
Length of introns and exons
Degradation of incorrectly spliced mRNA
Cleavage site of the terminator sequence and polyadenylation
Cleavage site of the terminator sequence and polyadenylation
Transport of mRNA through nuclear pore complex
Transcription of rRNA genes – tandem repeats
E. coli – 7 repeatsHuman – 200 repeatsXenopus – 600 repeats
Extensive rRNA processing
Nucleolus – the ribosome-producing factory
Functions of the nucleolus
Anatomy of the nucleus
Nuclear structures
From RNA to protein – the genetic code
Codon and anti-codon (tRNA)
Wobble base-pairing
Codon usageBacteria - 31 tRNAs/61 codons usedHuman – 497 tRNA genes/48 anticodons
Unusual nucleotides in tRNA
A
H
OH
Amino acid activation
Importance of amino acid activation
Proofreading by tRNA synthetases
Procaryotic and eucaryotic ribosomes
Structure of the rRNA components of the ribosome from E. coli
Structure of protein components of the ribosome from E. coli
tRNA binding sites at the ribosome
Initiation of protein synthesis in procaryotes
Shine-Delgarno sequence
Initiation of protein synthesis in eucaryotes
The translation cycle - movies
The translation cycle
Structure of the ribosome‘s active site
Active site of peptidyl synthase – the ribozyme
tmRNA (363b) provides ribosome rescue mechanism
Translational recoding - Selenocysteine incorporation
Translational recoding – frameshifting in retroviral RNA
Integrase – Pol, capsid protein - Gag
Formation of a mature protein
Molecular chaperone (hsp70)
Chaperonines – GroEL, GroES
Monitoring of protein quality after protein synthesis
20S
19S
Ubiquitin-conjugation marks protein for degradation
76 aa
Meaning of ubiquitin marks
Degradation of ubiquitinylated proteins in proteasome
Protein misfolding – example of PrP and PrP*prion disease
SummaryThe production of a protein by eucaryotic cell
RNA World and Evolution
Self-catalysis of RNA and its evolution
p-RNA: pyranosyl RNAPNA: peptide nucleic acid
Common elements of single-stranded RNA
In vitro isolation of a synthetic ribozyme – an example
Evolution from the RNA world