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CMSC 828N Introduction: Molecular biology background Class web page http://cbcb.umd.edu/confcour/CMSC828N.shtml 2 Course grading • 3 laboratory assignments • 15% (Labs 1,2,3) • Lab 1 given out by Sept 9, due Sept 23 (see syllabus) • Labs due by midnight on due date • Late penalty: 10%/day for 2 days maximum • 1 class presentation of a research paper • 5% • Lab 4 (mini-project) • 25% • Final exam • 25% 3 4 UMD Plagiarism policy Does this really happen? • Yes • Acknowledgement: many of the slides that follow are from Michael Brent, a professor at Washington Univ. What if I have a question? • You are required to ask if: – you have any doubt about whether or not you can use any (text,code,data) as part of your work for this class 5 Life Categories • Cellular organisms, viruses, prions • Cells are surrounded by a membrane –By weight, inside is mostly water –Generally, outside is aqueous, also • Major categories of cellular organisms are: –Prokaryotes –Eukaryotes 6 Prokaryotes • Single-celled organisms • Only 1 membrane. I.e., single compartment • Typically about 1 micron diameter 7 Prokaryotes 8 Eukaryotes • Single-celled organisms, plants & animals • Typical cell is 10 microns across (variable) • Membrane-bound nucleus contains DNA 9 Eukaryotes 10 Some other clades Tree of life: animals Entrez: taxonomy 11 Cell contents: small molecules Examples: • • • • Ions (Ca+, K+, Na+, Cl-) Sugars Fats Vitamins Can be obtained by • Import through membrane • Synthesis from imported precursors • Synthesis de novo 12 Macromolecules (polymers) Synthesis • Made in cell by linking monomers from a specified set Examples • Polysaccharides (sugar chains) • Proteins (amino acid chains) • DNA & RNA (Nucleic acids; nucleotide chains) 13 14 Protein functions Structural: e.g., • Cytoskeleton gives membrane strength & rigidity Signaling (information transduction) • receptors on cell surface sense hormones • DNA binding to turn genes on and off Enzymatic: speed up reactions to, e.g., • • • • Extract energy from nutrients Interconvert small molecules Immune response: bind and degrade invaders Maintain circadian rhythm & other clocks 15 Protein functions Enzymatic functions (cont.) • Programmed cell death (apoptosis) • Build macromolecular chains –Copy cell’s DNA during replication –Build other proteins from DNA instructions • Active transport through membrane –E.g. specific sugar transporters • Etc., etc., etc. Web resources • GO Browser, KEGG pathways, BioCarta pathways 16 Proteins • Built from 20 monomers called amino acids • Spontaneously fold into conformations determined by their amino acid sequences –Folded shape is essential to function • Often associate into complexes 17 Nucleic acids Two major types of nucleic acid polymers • Deoxyribonucleic acid (DNA) • Ribonucleic acid (RNA). Composition • Four monomers called nucleotides • DNA: deoxy – Adenine (A), Guanine (G), Cytosine (C), Thymine (T) • RNA: – Adenine (A), Guanine (G), Cytosine (C), Uracil (U) 18 DNA Function: • Long term information storage & transmission Structure: • Normally, double-helix –Twisted ribbon Base pairing • A:T and G:C 19 20 2 strands of DNA Orientation • Every (D/R)NA chain has a 5’ and a 3’ end –Position of free attachment pt in sugar • Many biological processes go from 5’ to 3’ –Elongation: nucleotides added to 3’ end –Read-out: DNA->RNA->protein 21 Orientation & the double helix Double helix is “anti-parallel” • 5’ end of each strand at 3’ end of the other • 5’ to 3’ motion in one strand is 3’ to 5’ in the other Double helix has no orientation • Biology has no “forward” and “reverse” strand • Both strands are equal • Relative to any single strand, there is a “reverse complement” or “reverse strand” 5’TTTTACAGGACCATG 3’ 3’AAAATGTCCTGGTAC 5’ 5’CATGGTCCTGTAAAA 3’ 22 RNA • Normally single-stranded • Much less stable than DNA. Shorter lifetime. • Can form complex structure by self-base-pairing 23 RNA self-base-pairing 24 DNA mRNA Protein • RNA polymerase transcribes a segment of DNA to a complementary messenger RNA • In eukaryotic cells: – Primary messenger RNA is processed to create mature mRNA – this processing involves splicing out certain segments of the RNA called introns – mature mRNA then transported out of the nucleus • Mature mRNA is translated into protein – by a ribosome 25 3D shape of transfer RNA 26 Quicktime animation 27 RNA Processing 28 RNA splicing • Splice sites are encoded in the sequence. • Splice site recognition is complex and imperfect. 29 Splice sites 30 Gene structure • Genes are highly structured regions of DNA • that ultimately yield a strand of amino acids 31 Translation of mRNA to Protein • DNA & mRNA represent protein sequences via a 3-letter code • there are 3 possible reading frames 32 Translation of mRNA to Protein • Each triplet is called a codon • The code is degenerate –61 codons map to 20 amino acids –Between 1 and 6 codons per amino acid –3 codons stop translation (TAA, TGA, TAG) –Codons for the same amino acid are called synonymous –DNA mutations that do not change the amino acid are called silent 33 34 Fun animations Quicktime animation: Protein synthesis Quicktime Animation: mRNA life cycle 35 Non-coding RNA Functions • Transfer RNAs: codon-to-amino-acid adapters • Ribosomes catalyze amino acid linkage –Protein-RNA complex. RNA is catalytic! • Small RNAs editing specific mRNAs, or • Prevent translation of specific mRNAs • All transcribed from DNA but not translated Structure • Shape, determined by self-pairing, is essential • External base-pairing is usually essential, too 36 Genes Molecular definition • Regions of DNA that are transcribed into a single RNA strand, with nearby DNA regions controlling time and quantity of transcription • Protein-coding genes and ncRNA genes Classical definition • Whatever it is that gives rise to a heritable trait 37 DNA Packaging • DNA is packed hierarchically • The chromosome is the largest package –Width: ~50 times that of smallest transistor –Humans have 22 chrs + 2 sex chrs • Human genome 1-2m long: 0.34nm/base • DNA is ~1 picogram (10-12g) per gigabase 38 Genome sizes • Widely varied • Not well correlated with organism complexity/sophistication –Typical bacterium: 1-10 megabases (mb) –Typical single-celled eukaryote: 10-30 mb –Smallest plants and animals: 100 mb (fruit fly, worm, mustard weed) –Human: 3 gb; some rats & gophers: 5-6 gb –Pine tree 60 g; Fern is 160 gb 39 ----------------------------------------------------------Organism Genome size ----------------------------------------------------------Amoeba dubia 670,000,000,000 Amoeba proteus 290,000,000,000 Ophioglossum petiolatum 160,000,000,000 Protopterus aethiopicus 139,000,000,000 Lilium longiflorum 90,000,000,000 Pinus resinosa 68,000,000,000 Lilium formosanum 36,000,000,000 Paramecium caudatum 8,600,000,000 Tarsius syrichta 5,151,600,000 Cercopithecus cephus 5,141,700,000 Zea mays 5,000,000,000 Hordeum vulgare 5,000,000,000 Macropus robustus 4,396,600,000 Parameles gunni 4,357,200,000 Monodelphis dimidiata 4,115,400,000 Pongo pygmaeus 4,046,300,000 Gerbillus pyramidum 3,913,100,000 Cercopithecus aethiops tantalus 3,898,300,000 Galago alleni 3,878,500,000 Didelphis marsupialis aurita 3,848,900,000 Ctenomys conoveri 3,848,900,000 Cebus capucinus 3,829,200,000 Ctenomys leucodon 3,824,200,000 Nicotiana tabaccum 3,800,000,000 Pan troglodytes 3,799,600,000 40