The Central Dogma of Molecular Biology

by

E. Börje Lindström

This learning object has been funded by the European Commissions FP6 BioMinE project

The flow of information

DNA molecule• General structure: - double stranded

- complementary

- helical

- antiparallel

• Strands: - backbone of alternating phosphate and

deoxyribos units

- four different bases; adenine (A), guanine (G), cytosine ( C ), and thymine (T).

• Double helix: - due to base pairing: A=T and GC

• Major and Minor groove

DNA molecule, cont.

• Size: - units: kilobase (kb) or kilobase pairs (kb pairs)

- E. coli chromosome 4 700 kb pairs

• Form: - closed chromosome molecule (in bacteria)

- 1 mm long packing problem in bacteria

- solved by supercoiling

• DNA binding proteins:

Un-specific: - histones

Specific: -Repressors

- RNA polymerase

- restriction enzymes

- modification enzymes

DNA molecule, cont.

DNA molecule, cont.

DNA replication

• Semi conservative: -new DNA molecules contain:

1 old strand and

1 new strand

• use a ’template’: - one of the strand is used

• ’primers’: -usually a piece of RNA

- DNA-polymerase unable to start replication

General

Initiation of replication• Start point: -only one (1) on the chromosome (300 bp)

- origin (ori)

ori

• bidirectional: - both directions

Synthesis of DNA (replication)• several enzymes involved (~ 20 pc)

- DNA helicase Unwinding the molecule

- DNA gyrase (topoisemerase II)

Open up (cut) the strands

- DNA-binding enzymes Protect ss-DNA from nucleases

- Primase Synthesises the RNA primer

- DNA-plymerase III Synthesis in direction 5’3’

There are 3 enz. in E. coli; pol I, II and III

- DNA-plymerase I Removes the primer

Repair any missing bp in DNA

- DNA ligase Makes a phospho-di-ester bond (glueing)

Synthesis of DNA, cont.• ’leading’ and ’lagging’ strands:

- leading: continous synthesis

- lagging: dis-continous synthesis

• proof-reading: - checking if any mitakes has been made

- pol. III removes the wrong nucleotides (3’ 5’)

Figures, DNA replication

RNA transcriptionThree types of RNA: • mRNA (genetical)

• tRNA (aa-carrier)

• rRNA (structural)

Structure: -ss-stranded (internal ds secundary structures)

- ribose

- four different bases; adenine (A), guanine (G), cytosine ( C ), and uracile (U).

Synthesis of RNA• ds DNA is the template: - only one of the strands

• RNA polymerase: - consists of four different subunits

- 2’

- 2’ = core enzyme

- recognises the start site

• Direction of synthesis: - 3’5’

Start and stop of RNA synthesis• Where is the start ?

- Note! No primers necessary!

- The polymerase binds to the promoter

- recognises and attaches to the promoter region

- ds-DNA opens up and the synthesis starts

- is detached and the core enzyme continues

• Where does the synthesis stop?

-termination at special DNA-sequenses, terminators

- inverted repeates in DNA ’stem-loop’-structures in RNA

PromotersA sequence in DNA upstreams a structural gene:

• -10 sequence Pribnow box

• Strong promoters bind effective

SG

-35bp -10bp

P

mRNA

• Short half-time

• Polycistronic (in bacteria) - information from several structural genes

• Definitions:

- operator (O): a gene that can be effected by a repressor protein

- operon: structural genes with the same repressor

SG1OP SG2 SG3

TranslationNecessary substances:

• mRNA

• ribosomes

• tRNA + aa tRNAaa (attached aa)

• different factors

• enzymes

• energy

tRNA

• DNA-genes: - Linear tRNA form (primary)

- cloverleaf structure (secundary)

• Two peoperties: - binds aa (enzymatic)

- binds to mRNA (codon) with its anti-codon

tRNA, cont.

Synthesis of proteinsA four (4) step process: • Initiation

• Elongation

• Termination-release

• Peptide folding

• Initiation: -a complex of

- 30S subunit,

- f-meth-tRNA, (start codon AUG in mRNA)

- mRNA and

- initiation factors are formed

• Shine-Delgarno sequence -3-9 bases in mRNA

- complementary to 16S rRNA

- addition of 50S subunit

Synthesis of proteins, cont.• Elongation: -several elongation factors are needed

- Next aa-tRNA is added to the A-site (ribosome)

- a peptide bond is created

- the peptide is moved to the A-site

- translocation to the P-site during

- movement of the ribosome forward

- a free A-site is created …

-Etc.

• polysomes: - mRNA with several ribosomes

Synthesis of proteins, cont.• Termination: -stop codes in mRNA

- UAA, UAG and UGA; nonsence codes

- no tRNA for these codes exist

- release factors RF1-3 release the protein

- the ribosomes disintegrate

• The genetic code: - in mRNA

• 3 bases - 1 aa

• 43 = combinations -but only 24 aa

- degenerated code

- the aa has several codes

Reading frame• Open reading frame (ORF): - a gene

AUG UAGS D-G

• Codon usage: -The code (tripletts) does not mean the same in all organisms

- The mRNA or ORF give different products

The wobble concept