Biomolecules: lecture 6

Transcription

1 Biomolecules: lecture 6 - to learn the basics on how DNA serves to make RNA = transcription - to learn how the genetic code instructs protein synthesis - to learn the basics on how proteins are synthesized = translation - to make friends with the biomolecules involved in these

2 Warming up for the day s theme: in terms of copying DNA, what is basically very different between replication (synthesis of new DNA from DNA) and transcription (synthesis of RNA from DNA)? replication transcription

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4 - DNA transcription reaction: (compare to DNA replication we looked at last time!)

5 - transcription and translation: which molecules are needed and what happens Transcription Translation DNA mrna protein mrna processing Amino acids Ribosomes Amino acid-specific transfer RNA s Folding & post-translational processing & trafficking

6 Translation = protein synthesis in nutshell: 20 different amino acids + lähetti-rna ribosomaalista RNA:ta osana ribosomin rakennetta over 20 different transfer RNA s (at least one for each amino acid) siirtäjä-rna Voet & Voet, Biochemistry, 4th edition

7 - ribosome is the factory - ribosome is built from protein and ribosomal RNA (rrna) - messenger RNA (mrna) is the code - transfer in nutsell: RNA (trna) brings in amino acids 20 different amino acids + lähetti-rna ribosomaalista RNA:ta osana ribosomin rakennetta over 20 different transfer RNA s (at least one for each amino acid) siirtäjä-rna Voet & Voet, Biochemistry, 4th edition

8 where do each reaction take place? nucleus cytosol

9 - there are 3 polynucleotide strands relevant in transcription: the 2 strands in double helical DNA, and the one RNA strand to be synthesized as the template for protein synthesis coding DNA = koodaava nauha >> the strand, whose sequence will be copied sense template DNA = mallinauha >>the strand, which is actually used in the copying antisense mrna = messenger RNA = lähetti-rna >> a copy of the coding strand sense

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11 - the transcription start place (initiation) in the gene: the promoter (promootteri) - E. coli promoters for different E. coli genes are very similar, and can be recognized box and -10 box - RNA polymerase binding site! upstream downstream tyypillinen operon = gene(s) under the control of one promoter

12 - prokaryotic and eukaryotic promoters are different

13 - a stem-loop structure, which will form in the mrna, terminates transcription Please notice how/why the stem loop can form!

14 Three types of RNA relevant in translation Messenger (mrna) lähetti - Template for protein synthesis - up to 5000 nucleotides long Ribosomal (rrna) ribosomaalinen - Structural component of ribosomes -120 to 3,700 nucleotides long Transfer (trna) siirtäjä - Carries amino acid to ribosome, decoder - around 75 nucleotides long - there are also additional RNA species in cells (but more of them on later courses)

15 The various RNA molecules relevant in protein synthesis ribosomal RNA structural component of ribosomes messenger RNA contains the information from DNA interface view of H. marismortui 50S ribosomal subunit. RNA is in gray and various proteins in other colors. Note the amount of RNA. Voet & Voet, Biochemistry, 4th edition transfer RNA brings the amino acid to the ribosome

16 Why Uracil in RNA and Thymine in DNA? O N NH 2 O H N O H N H H Deamination H N H H CYTOSINE (C) URACIL (U) Uracil base pairs with adenine not guanine => this reaction is mutagenic. Repair system recognises uracil as foreign to DNA

17 mrna gets modified after transcription cap prevents breaking up the mrna - what is the practical advantage for the cell synthesizing the protein? tail stabilizes the mrna i.e. works for the same purpose as the cap + also is used as an identification

18 Intron-splicing intron = non-coding sequence of DNA in-between coding regions in eukaryotic genes splicing (silmukointi) = removal of introns to create a mature (valmis) mrna

19 also trna gets modified after transcription - less well understood why this happens

20 Translation a codon triplet an amino acid

21 Each amino acid is coded for by three bases (codon) Why three? There are 4 different bases and twenty different amino acids 1 base code => 4 combinations 2 base code => 4 2 = 16 combinations 3 base code => 4 3 = 64 combinations There are specific codons for starting translation: AUG stopping translation: UAG UGA UAA The code is degenerate i.e. there is more than one codon per amino acid The genetic code is nearly universal for animals, plants and bacteria

22 Initiation of translation

23 base triplets in translation: see next page Let s play around with the importance of each base in a triplet!! (write here your short notes)

24 1st 2nd 3rd U C A G UUU Phe UCU Ser UAU Tyr UGU Cys U U UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA stop UGA stop A UUT UUG Leu UCT UCG Ser UAG stop UGG Trp G CUU Leu CCU Pro CAU His CGU Arg U C CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G AUU Ile ACU Thr AAU Asn AGU Ser U A AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G GUU Val GCU Ala GAU Asp GGU Gly U G GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G

25 1st 2nd 3rd U C A G UUU Phe UCU Ser UAU Tyr UGU Cys U U UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA stop UGA stop A UUT UUG Leu UCT UCG Ser UAG stop UGG Trp G CUU Leu CCU Pro CAU His CGU Arg U C CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G AUU Ile ACU Thr AAU Asn AGU Ser U A AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G GUU Val GCU Ala GAU Asp GGU Gly U G GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G

26 1st 2nd 3rd U C A G UUU Phe UCU Ser UAU Tyr UGU Cys U U UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA stop UGA stop A UUT UUG Leu UCT UCG Ser UAG stop UGG Trp G CUU Leu CCU Pro CAU His CGU Arg U C CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G AUU Ile ACU Thr AAU Asn AGU Ser U A AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G GUU Val GCU Ala GAU Asp GGU Gly U G GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G

27 1st 2nd 3rd U C A G UUU Phe UCU Ser UAU Tyr UGU Cys U U UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA stop UGA stop A UUT UUG Leu UCT UCG Ser UAG stop UGG Trp G CUU Leu CCU Pro CAU His CGU Arg U C CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G AUU Ile ACU Thr AAU Asn AGU Ser U A AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G GUU Val GCU Ala GAU Asp GGU Gly U G GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G

28 1st 2nd 3rd U C A G UUU Phe UCU Ser UAU Tyr UGU Cys U U UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA stop UGA stop A UUT UUG Leu UCT UCG Ser UAG stop UGG Trp G CUU Leu CCU Pro CAU His CGU Arg U C CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G AUU Ile ACU Thr AAU Asn AGU Ser U A AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G GUU Val GCU Ala GAU Asp GGU Gly U G GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G

29 1st 2nd 3rd U C A G UUU Phe UCU Ser UAU Tyr UGU Cys U U UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA stop UGA stop A UUT UUG Leu UCT UCG Ser UAG stop UGG Trp G CUU Leu CCU Pro CAU His CGU Arg U C CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G AUU Ile ACU Thr AAU Asn AGU Ser U A AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G GUU Val GCU Ala GAU Asp GGU Gly U G GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G

30 The universal genetic code is not really universal Codon Standard Mitochondrial UGA Stop Trp AUA Ile Met AGA Arg Stop AGG Arg Stop - the genetic code is, however, almost universal - mrna of one species can be translated by the protein synthesis machinery of another species What does the last fact immediately suggests?

31 Ribosomes Ribosome Small subunit Large subunit Sedimentation coefficient 70S 30S 50S Mass (kd) RNA 16S, 1542 nucleotides 23S, 2904 nucleotides 5S, 120 nucleotides RNA mass (kd) Proportion of mass 66% 60% 70% Number of proteins Protein mass (kd) Data from E.coli. Eukaryotic ribosomes are larger and more complex

32 Brief introduction of the molecules ribosomes - large protein complex - small and large subunit - contains also RNA Mathews et al. Biochemistry, 4th edition

33 trna

34 Translation = protein synthesis in nutshell: 20 different amino acids + over 20 different transfer RNA s (at least one for each amino acid) Voet & Voet, Biochemistry, 4th edition

35 - mrna exits from the nucleus to the cytoplasm - ribosomes read the mrna code and synthesize the protein - protein folds (laskostuu) into an active form

36 Charging of a trna with an amino acid (aminohapon kiinnittäminen spesifiseen siirtäjä-rna:han) (= jokaisella aminohapolla omansa!) This reaction is catalysed by an aminoacyl trna sythetase 3 3

37 Protein Forces involved in DNA structure 1. Covalent bonds 2. Short range repulsions 3. Electrostatic forces 4. Dipole-dipole interactions 5. Van der Waals forces