Week6 The molecular basis for growth and reproduc9on

Transcription

1 Week6 The molecular basis for growth and reproduc9on 1. Review: Central dogma (and beyond) 2. Structure of DNA and its replica9on 3. Transla9ng the gene9c code 4. Using ribosomal RNA to infer the evolu9onary rela9onships

2 DNA polymerase RNA polymerase Ribosome, mrna, trna folding spontaneous; aided by molecular chaperons

3 Many interes9ng processes occur beyond the central dogma Ribozymes (i.e. RNA enzymes) Post- transla9onal protein modifica9on (e.g. cell signaling, cancer) Prions (e.g. in BSE) Non- ribosomal pep9des (e.g. cyclosporine) RNA interference (i.e. RNA- mediated gene silencing) Reverse Transcriptase (e.g. HIV) RNA modifica9on Epigene9cs (i.e. modifica9on of DNA packing) Splicing Transposable elements

4 Gene9c informa9on, the structure of DNA A pairs T, G pairs C (hydrogen bonds) phosphate- sugar backbone (nega9ve charge) The two strands run an9parallel Double helix X- ray fiber diffrac9on by Rosalind Franklin Crick & Watson: The secret of life right handed chirality Semi- conserva9ve copying - > Homework paper

5 Example: Transla9ng the gene9c code Rules of transla9ng the code: - Non- overlapping triplet code - Redundant - 1 start and 3 stop signals 4 nucleo9des, 20 amino acids e.g. DNA: 5 -TGCGCCCTTACAGAATGCCATTGA-3! mrna:!5 -UGCGCCCUUACAGAAUGCCAUUGA-3! Protein: N -C A L T E C H *-C!

6 The ribosome translates the gene9c code into protein S: Svedberg unit of sedimenta9on Bacteria, Archaea rrna sequences are highly conserved - > evolu9onary rela9onship between species

7 The small ribosomal subunit reads the mrna, the large subunit synthesizes the protein X X makes pep(de bond X selects correct trna X MOVIE

8 Protein synthesis by the ribosome

9 Example: The 16S/18S RNA sequence is highly conserved between species Region decoding the mrna Eucarya (18S) Bacteria (16S) Archaea (16S) - The core of the RNA molecules is highly conserved - Differences can be used to infer evolu9onary rela9onships ( tree of life )

10 Rela9onship between structure and func9on in proteins GWQAEIVTEFSLL NEMVDVDPQGIL KCVDGR Primary structure: Sequence of amino acids connected by pep9de bonds α β hydrogen bonds formed by the protein backbone Secondary structure: Sequence of structural mo9fs formed by regular hydrogen bonds of the pep9de bonds (alpha helix, beta sheet) Ter9ary structure: 3D atomic model of the protein including cofactors (e.g. ions). Quaternary structure: Molecular assemblies spacefill representa9on cartoon representa9on e.g. ribosomes

11 The end

12 Why is 16S RNA highly conserved in all species? View from side Region where pep9de bond is made Small subunit (mainly 16S RNA) Region where correct trna is detected When the correct trna is binding to the mrna codon, 16S rrna closes and triggers pep9de bond forma9on

13 Week8: Obtaining the the 16S RNA sequence of a bacterium Sample - > DNA isola9on (Proper9es: DNA nega9ve, large; remove RNA with RNase) - > Amplifica9on of the 16S RNA gene (PCR) - > DNA sequencing

14 Polymerase chain reac9on (PCR) Key insights (1960- early 80s): DNA helix separated when temperature increases DNA polymerase copies DNA at lower temperature Heat- stable DNA polymerase discovered Yellowstone

15 The ribosome orchestrates protein synthesis using mrna, trna and addi9onal proteins (transla9on factors) 4 main phases of transla9on: 1. Ini9a9on 2. Elonga9on 3. Release 4. Recycling MOVIE

16 Chemistry and physics are the founda9on of biology Enzymes catalyze chemical reac9ons that are unfavorable. Physical techniques are used to visualize/measure biological processes. The tools of molecular biology, the combina9on of chemistry and gene9cs, have revolu9onized our ability to inves9gate and control cells.

17 Polymerase chain reac9on (PCR) Youtube Video - > Homework assignment: DNA replica9on

18 The ribosome as an example for a molecular machine 2 key func9ons: Amino Acid - select correct amino acid Transfer RNA (trna) adapter Messenger RNA (mrna) - form covalent bond between growing protein and next amino acid Transla9on involved many proteins and several large RNA molecules working together in a coordinated manner.