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

RNA folding

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

RNA splicing

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

Nucleolar fusion

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

The termination

A polyribosome

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

Protein folding

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

Ribozymes

In vitro isolation of a synthetic ribozyme – an example

Evolution from the RNA world