Bioinformatics Practical for Biochemists
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1 Bioinformatics Practical for Biochemists Andrei Lupas, Birte Höcker, Steffen Schmidt WS 2012/ DNA & Genomics 1
2 Description Lectures about general topics in Bioinformatics & History Tutorials will provide you with a toolbox of bioinformatics programs to analyze data Hands-On sessions will give you the opportunity to use these tools 2
3 Course Outline Mon Tue Wed Thr Fri DNA & Genomics Introduction to Proteins Annotation of Sequence Features Protein Classification Evolution & Design Course Material: eb.mpg.de/research/departments/protein-evolution/teaching 3
4 Course Outline 13:00-14:00 14:15-17:30 Presentation Tutorial (2 x 30min) & hands-on practical You will need to keep an electronic lab notebook Fri afternoon: Test Exercises 4
5 Software Requirements Browser (e.g. Firefox) Advanced Word Processor PyMOL ( free for teaching) 5
6 DNA & Genomics 1953 Model of DNA (F. Crick) 6
7 What is the genetic material? 1865 Gregor Mendel basic rules of heredity 1869 Friedrich Miescher discovery of nuclein (DNA), Hoppe-Seyler repeated all experiments 1881 Edward Zacharias chromosomes are composed of nuclein 1899 Richard Altmann renaming nuclein to nucleic acid wikipedia.org 7
8 DNA is the transforming material 1928 Frederick Griffith transforming principle - Str. pneumoniae experiment 1944 Avery & McCarty Griffith s transforming principle is DNA 8 history.nih.gov / wikipedia.org
9 DNA is the genetic material 1950 Erwin Chargaff A/T, C/G same amount in different tissues 1952 Hershey & Chase DNA is the genetic material using 32 P/ 35 S Phage/E. coli experiment 9 bacteriophagetherapy.info /
10 Solving the DNA structure 1952/53 Linus Pauling beat Cavendish Lab in discovery of α-helix Cavendish Lab (Cambridge) Watson & Crick allowed to work full-time on DNA Pauling shared manuscript with Cavendish Lab before publication (via his son Peter Pauling) 10
11 Solving the DNA structure 1952 Franklin & Wilkins X-ray of B-DNA - Wilkins showed results to Watson & Crick periodicity, phosphates are outside 1953 Crick & Watson model of B-DNA 11
12 Solving the DNA structure Nature,
13 DNA structure 13
14 Getting the code 1953 George E. Palade RNA organelles (ribosomes) 1957 Crick et.al suggest non-overlapping triplets only 20 out of 64 triplet code for an amino acid comma-free code 14
15 Getting the code 1961 Nirenberg & Matthaei polyu mrna produces polyf protein complete genetic code 1961 Sydney Brenner no overlapping codes concept of mrna triplet Code (Crick, Brenner, Barnett, Watts-Tobin) Starlinq point 3,, ;$I Overlappirq code +7 NUCLEIC ACID * I ---,-J+-~ ' Non-overlapplnq Code ETC. 15
16 Getting the code incl. start & stop codons Alternative start codon AUG (83%) GUG (14%) UUG (3%) Alternative stops UAA (63%, ochre ) UGA (29% opal ) / or Sec (Seleoncys) UAG (8%, amber ) E. coli 16
17 Gene Structure 1977 Sharp & Roberts pre-mrna is processed 1982 Cech ribo(nucleic en)zymes 1980 Joan A. Steitz role of snrnps in splicing wikipedia.org / yale.edu 17
18 Gene Structure Eurkayotes / Prokaryotes lac Operon 1: Regulatory gene 3: ß-galactosidase 4: ß-gal permease 8: ß-gal transacetylase Promotor region 18
19 Gene structure Polysomes in Prokaryotes EM picture of polysomes on a chromosome mrna with Ribosomes Transcription DNA initiation Miller, O. L. et al. Visualization of bacterial genes in action. Science 169,
20 Gene Structure Prokaryotic Operons lac Operon 1: Regulatory gene 3: ß-galactosidase 4: ß-gal permease 8: ß-gal transacetylase Promotor region 20 Griswold, A. (2008) Nature Education 1(1) Understanding Bioinformatics, Zvelebil & Baum, 2007
21 Gene Structure Prokayotes u-tokyo.ac.jp 21
22 Gene Structure Eurkayotes / Prokaryotes lac Operon 1: Regulatory gene 3: ß-galactosidase 4: ß-gal permease 8: ß-gal transacetylase Promotor region 22
23 Gene Structure Eukaryotes zazzle.com 23
24 Gene Structure Comparison! Often&have&introns& Eukaryote! Prokaryote! Genes! Gene!regulation! Repetitive!sequences! Organelle! (subgenomes)! Intraspecific&gene&order&and&number& generally&relatively&stable&& many&non8coding&(rna)&genes& There&is&NOT&generally&a&relationship& between&organism&complexity&and&gene& number& Promoters,&often&with&distal&long&range& enhancers/silencers,&mars,&transcriptional& domains& Generally&mono8cistronic& Generally&highly&repetitive&with&genome&wide& families&from&transposable&element& propagation& Mitochondrial&(all)& chloroplasts&(in&plants)& No&introns& Gene&order&and&number&may& vary&between&strains&of&a&species& Promoters& Enhancers/silencers&rare&& Genes&often®ulated&as& polycistronic&operons& Generally&few&repeated& sequences& Relatively&few&transposons& Absent& 24
25 Genomic era 1975 Frederick Sanger dideoxy sequencing 1986 Human Genome Initiative Genomes 1995 H. influenca 1.8 Mb 1.7k genes 1997 E. coli 4.6 Mb 4.3k genes 1996 S. cerevisiae 12.5 Mb 5.7k genes 1998 C. elegans 100 Mb 21.7k genes 2000 D. melanogaster 121 Mb 17k genes 25
26 Prokaryotic Genome E. coli 6 Mbp 1 by 2 µm cell size Kavanoff, Nature Education : Supercoiled chromosome of E. coli. 26
27 The human genome 2001 Draft H. sapiens 2.9 Bb 20-30k genes Science (2001), Nature (2001) 27
28 The human genome 28
29 Gene content 29
30 Genome Structure Comparison! Size! Eukaryote! Large&(10&Mb& &100,000&Mb)& There&is¬&generally&a& relationship&between&organism& complexity&and&its&genome&size& (many&plants&have&larger& genomes&than&human!)& Prokaryote! Generally&small&(<10&Mb;&most&<&5Mb)& Complexity&(as&measured&by&#&of&genes& and&metabolism)&generally&proportional& to&genome&size& Content! Most&DNA&is&nonLcoding& DNA&is& coding&gene&dense & Telomeres/! Centromeres! Number!of! chromosomes! Chromatin! Present&(Linear&DNA)& More&than&one,&(often)&including& those&discriminating&sexual& identity& Histone&bound&(which&serves&as&a& genome®ulation&point)& Circular&DNA,&doesn't&need&telomeres& Don t&have&mitosis,&hence,&no& centromeres.& Often&one,&sometimes&more,&Lbut& plasmids,¬&true&chromosome.& No&histones& Uses&supercoiling&to&pack&genome& & 30
31 Gene content 31
32 Human Genome Content Segmental duplications LTR retrotransposons DNA transposons Simple sequence repeats 5% 3% 2.9% 8.3% 13.1% SINEs Miscellaneous heterochromatin 8% 20.4% LINEs 11.6% 1.5% Miscellaneous unique sequences 25.9% Protein-coding genes Introns Gregory (2005), Nature 32
33 Gene Structure Eukaryotic Gene Mammal Cons Scale chr1: SMG5 4 _ 10 kb hg19 156,225, ,230, ,235, ,240, ,245, ,250,000 UCSC Genes (RefSeq, UniProt, CCDS, Rfam, trnas & Comparative Genomics) Placental Mammal Basewise Conservation by PhyloP -4 _ Common SNPs(135) RepeatMasker Simple Nucleotide Polymorphisms (dbsnp 135) Found in >= 1% of Samples Repeating Elements by RepeatMasker 33
34 Human Genome Content Segmental duplications LTR retrotransposons DNA transposons Simple sequence repeats 5% 3% 2.9% 8.3% 13.1% SINEs Miscellaneous heterochromatin 8% 20.4% LINEs 11.6% 1.5% Miscellaneous unique sequences 25.9% Protein-coding genes Introns Gregory (2005), Nature 34
35 Transposable Element - Mobile Elements / Jumping genes Barbara McClintock ( ) studies in the 40 s & 50 s of spotted kernels in maize discovery of controlling elements initially thought to be unique to maize but later also found in eukaryotes, bacteria, viruses, phages & plasmids Nobel prize in 1983 wikipedia.org 35
36 Transposable Element - Mobile Elements / Jumping genes DNA Transposons transposase cuts out transposon & inserts it at the target site cut-and-paste mechanism prokaryotes & eukaryotes Retrotransposons transposon DNA transcribed to RNA insertion to genome by reverse transcription LTR, LINEs, SINEs eukaryotes only wikipedia.org 36
37 What can Bioinformatics do for you? sequence analysis comparison, annotation, phylogeny genomics assembly, gene finding / annotation, phylogeny data mining / analysis text mining, expression profiling (microarray, RNAseq), image analysis structural bioinformatic 2 nd ary structure prediction, protein design, docking 37
38 38
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