From DNA to Protein Class 4 02/11/04 RBIO-0002-U1
From DNA to Protein 02/11/04 RBIO-0002-U1
From DNA to Protein Genome DNA Expressed Sequence Transcriptome RNA Gene Genome DNA Expressed Sequence Transcriptome RNA Protein Proteome 02/11/04 RBIO-0002-U1
Transcription Begins with unwinding of the DNA One DNA strand is used as a template for RNA synthesis RNA polymerase synthesizes the transcript The transcript is complementary to the template The RNA chain is released from the DNA template and the DNA helix reforms 02/11/04 RBIO-0002-U1
RNA polymerase Catalyzes a similar reaction as DNA polymerase, but: Uses ribonucleotides Can start a chain without primer Has a higher error rate than DNA polymerase: 1 in 104 nucleotides vs 1 in 107 nucleotides 02/11/04 RBIO-0002-U1
RNA RNA can serve as structural and enzymatic component mRNA (messenger RNA) specifies the sequence of proteins tRNA (transfer RNA) forms adaptors between codon and amino acid rRNA (ribosomal RNA) is part of the ribosome snRNA (small nuclear RNA) is involved in splicing These are the functions we understand well enough to put in textbooks. http://www.yangene.com/content22_6.htm 02/11/04 RBIO-0002-U1
Transcription of a bacterial gene 02/11/04 RBIO-0002-U1
DNA OpenReadingFrame DNA Exon Intron Gene Coding Sequence Regulatory elements Termination sites Exon Intron polyA signal Promoter 02/11/04 RBIO-0002-U1
Transcription from DNA to RNA RNA Polymerase + Transcription factors Pre mRNA Cleavage Splicing Polyadenylation (Editing) mRNA AUG AAAAAAAAAAAAA(A)n 5’UTR ORF 3’UTR 02/11/04 RBIO-0002-U1
mRNA processing in Eukaryotes The primary transcript undergoes several processing steps in the nucleus A 5’ cap is added Introns (non coding sequences) are removed by splicing A polyA tail is added to the 3’ end The mature mRNA is exported to the cytoplasm, where protein synthesis takes place Only 5% of the transcribed RNA reaches the cytoplasm 02/11/04 RBIO-0002-U1
Prokaryotic and Eukaryotic Transcripts 02/11/04 RBIO-0002-U1
Splice sites Life is complicated: 02/11/04 RBIO-0002-U1 http://www.swbic.org/education/comp-bio/images/spliceconsensus.gif 02/11/04 RBIO-0002-U1
The Splicosome consists of snRNPs and other proteins snRNRs: small nuclear ribonucleoprotein particles 02/11/04 RBIO-0002-U1
Splice Variants 02/11/04 RBIO-0002-U1
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Ensembl Genome browser 02/11/04 RBIO-0002-U1
NCBI 02/11/04 RBIO-0002-U1 http://www.ncbi.nlm.nih.gov/ http://www.ncbi.nlm.nih.gov/LocusLink/ 02/11/04 RBIO-0002-U1
mRNA export 02/11/04 RBIO-0002-U1
RNA stability The lifetime of a mRNA can vary from minutes to hours The 3’ UTR is involved in RNA stability RNA stability allows to regulate the level of protein Fast degradation of RNAs provides a quick response mechanism 02/11/04 RBIO-0002-U1
Expression levels vary 02/11/04 RBIO-0002-U1
Road map: From DNA to Protein 02/11/04 RBIO-0002-U1
From RNA to Protein: The genetic code 4 Nucleotides have to code for 20 Amino acids Triplet code can specify 64 (4 x 4 x 4) amino acids The genetic code is redundant. Some amino acids are encoded by more than one triplet A triplet is called codon 02/11/04 RBIO-0002-U1
The Genetic Code 02/11/04 RBIO-0002-U1
The Genetic Code 02/11/04 RBIO-0002-U1
Translation: Reading Frames 02/11/04 RBIO-0002-U1
Example: Reading Frames A reading frame uninterrupted by Stop codons is called: Open Reading Frame (ORF) “SequencherTM” 02/11/04 RBIO-0002-U1
Transfer RNA (tRNA) recognizes and binds codon on one side, the matching amino acid on the other side 02/11/04 RBIO-0002-U1
tRNA and redundancy Some amino acids are specified by more than on codon There are more than one tRNA for some amino acids Some tRNAs tolerate mismatches at position 3 (wobble) 02/11/04 RBIO-0002-U1
Adaptors Aminoacyl-tRNA synthetase couples covalently amino acid and corresponding tRNA molecule There are 20 different synthetases, one for each amino acid. They recognize specific nucleotides in anticodon and accepting side 02/11/04 RBIO-0002-U1
The Ribosome Travels along mRNA “captures” tRNA Holds tRNA in position Forms bond There are millions of ribosomes per cell Several ribosomes can transcribe a mRNA at the same time Speed: 2 -20 aa / seconds 02/11/04 RBIO-0002-U1
Ribosomes in the cell 02/11/04 RBIO-0002-U1
Aminacyl-tRNA Peptidyl-tRNA Exit 02/11/04 RBIO-0002-U1
Elongation 02/11/04 RBIO-0002-U1
Initiation 02/11/04 RBIO-0002-U1
Termination 02/11/04 RBIO-0002-U1
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Websites http://www.blc.arizona.edu/INTERACTIVE/DNA3/proteins.html 02/11/04 RBIO-0002-U1
Translation from RNA to Protein AAAAAAAAAAAAA(A)n AUG Ribosome Protein M Proteolytic Cleavage Chemical Modification Folding 02/11/04 RBIO-0002-U1