Structural Components of Nucleotides 11 Base Sugar IntroductionNucleotide to Cells & Microscopy and Nucleic Acid Phosphate Structure Glycosidic bond H NUCLEOTIDE H Nucleic acid – polymer of nucleotides – directionality 5’à3’ When you write a sequence: ATCG It is assumed that the 5’-end is on the left and the 3’-end is on the right, unless otherwise labeled. Phosphodiester bonds RNA 5’-ATCG-3’ 3’-GCTA-5’ same molecule DNA Composition of DNA? Table 3-1 1 Chargaff’s Rules DNA is Double Stranded Helix http://higheredbcs.wiley.com/legacy/college/voet/0470129301/kinemages/exercise_2.html Computer-simulated space-filling model of DNA. Figure 3-8 2 Video: Computer-simulated space-filling model of DNA. • The crucial piece of evidence for DNA structure came from X-ray “crystallography.” Wilkins learned how to purify DNA and make regular fiber patterns. Rosalind Franklin performed the X-ray diffraction and deduced there was a helix. • Francis Crick saw the data at a seminar Wilkins gave and also deduced there was a helix and the size parameters. • James Watson discovered how the bases went together (complementarity) using Chargaff rules (A=T, G=C). • Watson & Crick published their structure in 1953. Beautiful example of how structure predicted function. SUMMARY 12 (34 Å) IntroductionCentral to Cells Dogma & Microscopy of Molecular Biology Right-handed, antiparallel, double- sugar–phosphate stranded helix. With the “base backbone complementarity,” it explains genetic material: (phosphodiester bonds) • Storage of genetic information • Replication • Information retrival 3 From DNA to Protein: Gene Expression Central Dogma The central dogma of molecular biology • Central Dogma: from Genes to Proteins Replication • Replication of the genes (DNAàDNA) • Transcribing the information (DNAàRNA) • Translating the nucleotide sequence into protein sequence (RNAàProtein) – The Genetic Code Information Flow – Protein Biosynthesis DNA replication is semiconservative (Meselson-Stahl Expt) Replication 4 DNA Replication Each New DNA Strand Grows from Its 5´ End to Its 3´ End Arthur Kornberg showed that DNA contains information for its own replication. He combined in a test tube: DNA, the four deoxyribonucleoside triphosphates (dNTPs–monomers), DNA polymerase, salts (Mg+2), and buffer. The DNA served as a template for synthesis of new DNA. Each New DNA Strand Grows from Its 5´ End to Its 3´ End ALL polymerases add nucleotides to the 3’ end (Direction is termed 5’ g 3’) Transcription Pyrophosphatase 34 5 Central Dogma Central Dogma The central dogma of molecular biology RNA is key to this process: Replication • Messenger RNA (mRNA)—carries copy of a DNA sequence to site of protein synthesis at the ribosome • Transfer RNA (tRNA)—carries amino Messenger RNA Transfer RNA acids for polypeptide assembly (mRNA) (tRNA) Ribosomal RNA • Ribosomal RNA (rRNA)—catalyzes (rRNA) peptide bond formation and provides structure for the ribosome Transcription Central Dogma Transcription components: The central dogma of molecular biology • A DNA template for base pairings—one of the two strands of DNA Replication • Nucleoside triphosphates (ATP,GTP,CTP,UTP) as substrates Transcription • An RNA polymerase enzyme Messenger RNA Transfer RNA Transcription process: (mRNA) (tRNA) • RNA polymerase unwinds DNA about ten base pairs at a Ribosomal RNA time; reads template in 3’ to 5’ direction, synthesizes RNA (rRNA) in the 5’ to 3’ direction. • The RNA transcript is antiparallel to the DNA template strand, and adds nucleotides to its 3’ end. • NTPs incorporate NMP and PPi is a product! 6 Transcription Transcription: Where to start? • Production of mRNA transcript by RNA polymerase 5' Flanking Coding Region 3'-flanking prokaryotes Promoter eukaryotes The consensus sequence for each element in human genes (N is any nucleotide) Transcription Translation 7 Central Dogma The central dogma of molecular biology Replication Translation Messenger RNA Transfer RNA (mRNA) (tRNA) Ribosomal RNA • The Code (rRNA) • The Adaptors (tRNA) • The Ribosome (rRNA + rProteins) What is the relationship between a DNA sequence and an amino acid sequence? Translation: The Genetic Code Translation: The Genetic Code The genetic code is redundant. The genetic code is universal. The genetic code: Specifies which amino acids will be used to build a protein Codon: A sequence of three bases—each codon specifies a particular amino acid. Start codon: AUG—initiation signal for translation. Stop codons: UAA, UAG, UGA—stop translation and polypeptide is released. 8 Translation: tRNA Translation: tRNA tRNAs must deliver amino acids corresponding to each codon The conformation (three-dimensional shape) of tRNA results from base pairing (hydrogen bonding) within the molecule. 3‘-end is the amino-acid attachment site—binds covalently. At the other end (middle of the tRNA sequence) is the Anticodon—site of base pairing with mRNA. Unique for each species of tRNA. Translation: Ribosome Translation: tRNA Ribosome: the workbench—holds Template for mRNA – mRNA and charged tRNAs in the read 3’à5’ correct positions to allow assembly of polypeptide chain. Ribosomes are not specific, they can tRNA anticodon make any type of protein. N C 9 Translation: Protein Biosynthesis: Ribosome Structure Ribosomes have two subunits, large and small. When not active in Translation: Ribosome translation, the subunits exist separately. • The small subunit (40S) has one ribosomal RNA (rRNA) (18S) and 33 proteins. • The large subunit (60S) has three molecules of rRNA (28S, 5.8S, 5S) and 49 different proteins. • Ribosomal subunits are held together by ionic and hydrophobic forces (not covalent bonds) (80S). Translation: Protein Biosynthesis; Elongation Translation: Protein Biosynthesis; Elongation ELONGATION EF-Tu GTP Decoding Translocation (GTP hydrolysis) (GTP hydrolysis) Peptidyltransferase 10 Central Dogma The central dogma of molecular biology Replication Animated videos of DNA structure and Central Dogma (https://wileyassets.s3.amazonaws.com/Voet_Fundamentals_of_Biochemistry_5e_ISBNEPROF12533/media/ Guided_Tour/dnaStructure.html) 11.