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 G  C Major and Minor groove

DNA molecule, cont. Size: - units: kilobase (kb) or kilobase pairs (kb pairs) - E. coli chromosome  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 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 helicaseUnwinding the molecule - DNA gyrase (topoisemerase II) Open up (cut) the strands - DNA-binding enzymesProtect ss-DNA from nucleases - PrimaseSynthesises 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 IRemoves the primer Repair any missing bp in DNA - DNA ligaseMakes 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 transcription Three 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

Promoters A sequence in DNA upstreams a structural gene: -10 sequencePribnow 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 SG 1 OPSG 2 SG 3

Translation Necessary substances: mRNA ribosomes tRNA + aa  tRNA aa (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 proteins A 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 4 3 = combinations -but only 24 aa - degenerated code - the aa has several codes

Reading frame Open reading frame (ORF): - a gene AUGUAG S 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