Regulation of gene expression III. tRNA synthesis, aminoacyl activation, translation, translation...

Regulation of gene expression III.

tRNA synthesis, aminoacyl activation, translation, translation inhibitors,

molecular genetics of deseases and diagnostic methods

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The genetic code


The structure of transfer RNA

Ψ: pseudouridine D: dihydrouridineDownloaded from www.pharmacy123.blogfa.com

Activation of aminoacids


Comparison of ribosomes


Initiation of translation

1. Peptidyl-transferase

2. EF-G (translocase)


Elongation

1. EF-Tu + GTP= EF-Tu-GDP + Pi

EF-Ts

EF-Tu + GDP

Important time delay for accuracy


Termination

Binding of releasing factor to STOP

Termination


Translation on the surface and processing in the lumen of ER


The recognition of signal peptide


Double transmembrane domain

Stop-transfer sequence

Start-transfer sequence


Multiple transmembrane domain


Differences in the pro-and eucaryotic mRNA


The procaryotic translation


Regulation of translation initiation

• dsRNA dependent protein kinase phosphorylates eIF, this inhibits initiation

• Viral interferon stimulated dsRNA dependent protein kinase (initial stage of infection:-RNA, +RNA or RNA:RNA hybrid)

• Reticulocyte protein kinase: inhibited by hem – no hem, no protein synthesis (90% globin)


The effect of antibiotics on procaryotic translation

• Steptomycin – inhibition of initiation, misreading of the code (resistance: mutation in the S12 ribosomal protein) Neomycin, gentamycin similarly

• Tetracycline – inhibits a. ac. tRNA binding to ribosome

• Chloramphenicol – inhibits pepidyl transferase


Eucaryotic translation specific inhibitors

• Puromycin – prevents tRNA binding to ribosome and incorporates into peptide chain, cause premature termination

• Cycloheximide – specific for eucaryotes, chloramphenicol-like effect

• Diptheria (corynebacterium exotoxin) the elF2 is inactivated by ADP-ribosilation made by the toxin


DNA chip

Green: less than average exp.

red: more than average exp.

black: close to average exp.

Differential analysis of mRNA levels of 1800 genes on cDNA array


Differential analysis of protein expression

pH2 pH12


Fluorescence-activated cell sorter


Sequencing I


Sequencing II.


Sequencing III.


RNA interference


Genetic diseases

• 4000 diseases have genetic base• 10% of hospitalised children• Many chronic diseases have genetic component

(diabetes melitus, athelosclerosis)• Chromosomal - trisomy of 21 – Down’s

syndrome (karyotype – chromosomal translocations – positional effect)

• Monogenic disorders (autosomal dominant – recessive – X-linked)

• Multifactorial dirorders (ishemic heart disease, hypertension)


Pedigree analysis

T to A change in the -globin gene – sickle cell anemia

SS homozygous

AS heterozygous (prenatal)

AA homozygous


Regulation of gene expression III. tRNA synthesis, aminoacyl activation, translation, translation...

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