BIO 344 1st EditionFinal Exam Study Guide Lectures: 1-23 Genome Structure- organization of genetic informationo know bacteria gene characteristics vs humansBacterial Genome- orf= frame of base pairs, begins with start codon, codes for a gene, and genes in one orf code for one protein- non protein coding genes= regulatory genes that code for tRNA- dense with coding sequence, functionally related genes are typically close together, can synthesize all amino acidsTrp Operon- operon= a unit made up of linked genes that regulates other genes responsible for protein synthesis- Trp operon= controls transcription of tryptophan- Restriction enzyme = enzyme that cleaves DNA at a specific locus of sequenceHuman Genome- definition of epigenetics- low gene density, not organized by function, transcription factors are combinationalDNA structure- antiparallel, polarity, base pairs held together by H- bonding- supercoils are formed to reduce strain of DNA unwinding in replication- positive supercoils vs negative supercoils- function of topoisomerase to relieve strain-- type I vs. Type II- gyrase present in bacteria since they do not have histonesReplication- initiation: know properties needed at the origin- DnaA, DnaB, and DnaC composition and function- Leading vs. lagging strand (Okazaki fragments)- Processivity= what keeps polymerase associated to the DNA strando Primase= low processivity, polymerase= high processivityo ss binding protein= coats single stranded DNA at fork to keep from tangling- Know problems with replication: okazaki fragments, cantenation, completing the endso Telomeres and telomeraseDNA polymerization- utilizes 3’ end, needs –OH to bind, overall synthesis 5’3’- know how cell differentiates between DNA and RNA- fidelity—exonuclease proofreading, mismatch repair- know how DNA polymerase are targets for chemotherapeutics- using radioactive and fluorescent labels to “see” DNADNA sequencing- know these methods: Sanger, Gel electrophoresis, PCR, NextGen- know Messelson Stahl experiment/paper on mechanisms of replicationGene Regulation- genomes have directed stable pathways—gene regulation is the mechanism for this- Lac Operon—metabolizing lactoseo Positive and negative control—need glucose absent and lactose presento Know role of beta galactosidase, X-gal, and that lactose is upstream from glucoseo Know complementation groupso Know which gene codes for beta galactosidase (lacY or lacZ) and how that information was obtained experimentally (end result is that its lacZ)o Know, experimentally, how we know how cells control their expression of their genes—how lacZ expression is altered in regulatory mutantso Constitutive vs. uninducible o lacI encodes repressor, trans working—know how this is known experimentallyo operator is cis working—know how this is known experimentallyStructure and Function of lac repressor - binding site is palindromic-- tells us lac repressor is a dimer- defining ligand binding domains—know what different mutations do- helix-turn-helix motif- mutations in DNA binding domain are dominantStructure and function of lac operator- O1 is a binding siteo Know, experimentally, how it is known that O2 and O3 also function in regulationDNA binding site—recognition of target- specificity and affinityo dimers boost both—know how- understand Kon, Koff, equilibrium constant and how to calculateLigand- induced allostery—when lactose is present- allolactose is an inducer- cooperativity- negative regulationIntegrating glucose and lactose signals- glucose repression is combinatorialo know effect of glucose and lactose presence, CAP bound? Operon on or off?o cAMP, CAP—know function of eachGeneralizing Regulation- relationships between negative/positive and inducible/repressible and effect on gene regulationGeneral Transcription- Know RNAP function, start site at +1, and consensus sequences and how they effect expression-- -35 and -10- RNA enzymeo Holoenzyme and core enzyme Know composition and differencesTranscription Initiation- stages initiated by RNAP- closed complex, open complex, abortive initiation, elongation complex- sigma factor melts the -10 regionEukaryotic Transcription- stem cell research—pluripotent cellso metastable statesEukaryotic Transcription--- RNAP- RNAP IIo Shaped similar to bacterial RNAPo Promoters, enhancers, and transcription factors Core promoter= start site- TATA box, DPA, initiator element Transcription factors—TFIID function and composition Enhancers- Tethering a protein to DNA- Combinatorial controlChromatin and Gene Expression- experiment with micrococcal nuclease digestion of DNA showed that something protectsour DNA- chromatin= DNA wrapped in nucleosomeso DNA wrapped around a histone octomer twice- Know histone structural components of the core and H1o Know function of H1o Histones are basic—Arg and Lys are positively charged and neutralize DNA to reduce electrostatic potential and the N-term carries a positive chargeo Allow for negative supercoiling in eukaryotes Understand this process Allows for access to DNA for replicationChromatin- 2 states: euchromatin (accessible) and heterochromatin (inaccessible)o Understand the conformation and implications of eachPromoter Accesso regions must be free of nucleosomeTranscriptional Activators in Chromatin- gal4-VP16= hybrid activator- histone modificationo acetylation by HATs and deacetylation by HDAC—know function and result loosen the grip of histones on DNA or tightens the grip, respectivelyRemodeling Complexes- three methods: sliding, acetylation, methylationo know the effects of eachHistone code- different modifications are recognized by proteins that perform different functions/ effect expressiono H3K9me silencing, H3K9ac expression- Protein complexes are readers and writers for the histone codeo Readers recognize residues Bromodomains and chromodomains (know which recognizes what)Activation- transcription factor GCN4 can recruit a coactivator (GCN5, a HAT)o weakens affinity for DNA- Kinase can phosphorylate H3S10 and can recruit TFIIDRepression- Histone methyl transferase (HMT)o Repress or silence transctiption- Histone demethylases (HDM)o Reverse methylationStress Induced epigenetic control of behavior- know Bdnf function and how it causes modification of geneso controlled by multiple promoters—know function of P4o know CHIP techniqueDNA methylation ( different from histone methylation)- DNMT, dMTase, SAM- Happens