The Central Dogma of Molecular Biology

byE. 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