BSC197 Lecture Notes 10.26 - 10.30
BSC197 Lecture Notes 10.26 - 10.30 BSC197
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Date Created: 10/31/15
BSC197 Lecture Notes 10262015 103015 1026 Lecture Overview The Flow of Genetic Information The information content of DNA is in the form of specific sequences of nucleotides The DNA inherited by an organism leads to specific traits by dictating the synthesis of proteins Proteins are the links between genotype and phenotype the process by which DNA directs protein synthesis includes two stages transcription and translation Basic Principles of Transcription and Translation RNA is the intermediate between genes and the proteins for which they code Transcription is the synthesis of RNA under the direction of DNA Transcription produces messenger RNA mRNA Translation is the synthesis of a polypeptide which occurs under the direction of mRNA Ribosomes are the sites of translation In prokaryotes mRNA produced by transcription is immediately translated without more processing In a eukaryotic cell the nuclear envelope separates transcription from translation Eukaryotic RNA transcripts are modified through RNA processing to yield finished mRNA A is the initial RNA transcript from any gene The central dogma is the concept that cells are governed by a cellular chain of command DNA 9 RNA 9 Protein The Genetic Code How are the instructions for assembling amino acids into proteins encoded into DNA There are 20 amino acids but there are only four nucleotide bases in DNA How many bases correspond to an amino acid Codons Triplets of Bases The ow of information from gene to protein is based on a triplet code a series of nonoverlapping threenucleotide words These triplets are the smallest units of uniform length that can code for all the amino acids Example AGT at a particular position on a DNA strand results in the placement of the amino acid serine at the corresponding position of the polypeptide to be produced During transcription one of the two DNA strands called the template strand provides a template for ordering the sequence of nucleotides in an RNA transcript During translation the mRNA base triplets called codons are read in the 5 to 3 direction Each codon specifies the amino acid to be places at the corresponding position along a polypeptide 0 Codons along an mRNA molecule are read by translation machinery in the 5 to 3 direction 0 Each codon specifies the addition of 20 amino acids Cracking the Code 0 All 64 codons were deciphered by the mid1960s 0 Of the 64 triplets 61 code for amino acids 3 triplets are the stop signals to end translation 0 The genetic code is redundanct but not ambiguous no codon specifies more than one amno acid 0 Codons must be read in the correct correct groupings in order for the specified polypeptide to be produced RNA polymerase Binding and Initiation of Transcription o Promoters signal the initiation of RNA synthesis 0 Transcription factors mediate the binding of RNA polymerase and the initiation of transcription 0 The completed assembly of transcription factors and RNA polymerase II bound to a promoter is called a transcription initiation complex 0 A promoter called TATA box is a crucial in forming the initiation complex in eukaryotes Elongation of the RNA strand 0 As RNA polymerase moves along the DNA it untwists the double helix 10 to 20 bases at a time o Transcription progresses at a rate of 40 nucleotides per second in eukaryotes 0 A gene can be transcribed simultaneously by several RNA polymerases Termination of Transcription dependent on loops 0 The mechanisms of termination are different in bacteria and eukaryotes 0 In bacteria the polymerase stops transcription at the end of the terminator 0 In eukaryotes the polymerase continues transcription after the premRNA is cleaved from the growing RNA chain the polymerase eventually falls off the DNA Eukarvotic cells Modify RNA after transcription 0 Enzymes in the eukaryotic nucleus modify premRNA before the genetic messages are dispatched to the cytoplasm 0 During RNA processing both ends of the primary transcript are usually altered 0 Also usually some interior parts of the molecule are cut out and the others are spliced together Alteration of mRNA ends 0 Each end of a premRNA molecule is modified in a particular way 0 The 5 end receives a modi ed nucleotide 5 cap 0 The 3 end gets a polyA tail 0 Opposite ends of the molecule 0 These modifications share several functions 0 They seem to facilitate the export of mRNA 0 They protect mRNA from hydrolytic enzymes 0 They help ribosomes attach to the 5 end 10282015 Lecture RNA Splicing 0 Removal of unneeded sequences 0 Most eukaryotic genes and their RNA transcripts have long noncoding stretches of nucleotides that lie between coding regions 0 These noncoding regions are called intervening sequences or intervening regions these are removed 0 The other regions are called meaning expressed because they are eventually expressed usually translated into amino acid sequences 0 removes introns and joins exons creating an mRNA molecule with a continuous coding sequence Molecular Components of Translation 0 A cell translates an mRNA message into protein with the help of 0 Molecules of tRNA are not identical 0 Each carries a specific amino acid on one end 0 Each has an on the other end the anticodon basepairs with a complementary codon on mRNA The Structure and Function of Transfer RNA 0 A tRNA molecule consists of a single RNA strand that is only about 80 nucleotides long 0 Flattened into one plane to reveal its base pairing a tRNA molecule looks like a cloverleaf 0 Accurate translation requires two steps 0 First a correct match between tRNA and an amino acid done by the enzyme 0 Second a correct match between the tRNA anticodon and an mRNA codon 0 Flexible pairing at the third base of a codon is called and allows some tRNAs to bind to more than one codon Ribosomes 0 Ribosomes facilitate specific coupling of tRNA anticodons with mRNA codons in protein synthesis 0 The two ribosomal subunits large and small are made of proteins and ribosomal RNA rRNA 0 A ribosome has three binding sites for tRNA 0 The P site polypeptide holds the tRNA that carries the growing polypeptide chain 0 The A site amino acid holds the tRNA that carries the next amino acid to be added to the chain 0 The E site is the exit site where discharged tRNAs leave the ribosome Building a Polypeptide o The three stages of translation 0 Initiation o Elongation 0 Termination o All three stages require protein factors that aid in the translation process Ribosome Association and Initiation of Translation 0 The initiation stage of translation brings together mRNA a tRNA With the first amino acid and the two ribosomal subunits 0 First a small ribosomal subunit binds With mRNA and a special initiator tRNA 0 Then the small subunit moves along the mRNA until it reaches the start codon AUG 0 Proteins called initiation factors bring in the large subunit that completes the translation initiation complex Elongation of the Polypeptide Chain 0 During the elongation stage amino acids are added one by one to the preceding amino acid 0 Each addition involves proteins called elongation factors and occurs in three steps codon recognition peptide bond formation and translocation Termination of Translation 0 Termination occurs When a stop codon in the mRNA reaches the A site of the ribosome 0 The A site accepts a protein called a release factor 0 The release factor causes the addition of a water molecule instead of an amino acid 0 This reaction releases the polypeptide and the translation assembly then comes apart Polyribosomes 0 A number of ribosomes can translate a single mRNA simultaneously forming a single 0 Polyribosomes enable a cell to make many copies of a polypeptide very quickly Completing and Targeting the Functional Protein 0 Often translation is not sufficient to make a functional protein 0 Polypeptide chains are modified after translation 0 Completed proteins are targeted to specific sites in the cell Point Mutations can affect Protein Structure and Function 0 are changes in the genetic material of a cell or virus are chemical changes in just one base pair of a gene 0 The change of a single nucleotide in a DNA template strand can lead to the production of an abnormal protein 10302015 Lecture Substitutions o A replaces one nucleotide and its partner with another pair of nucleotides 0 Silent mutations have no effect on the amino acid produced by a codon because of redundancy in the genetic code still code for amino acid but not necessarily the right amino acid change an amino acid codon into a stop codon nearly always leading to a nonfunctional protein Insertions and Deletions 0 and are additions or losses of nucleotide pairs in a gene 0 These mutations have a disastrous effect on the resulting protein more often than substitutions do 0 Insertion or deletion of nucleotides may alter a reading frame producing a What is a Gene Revisiting the Question 0 The idea of the gene itself is a unifying concept of life 0 We have considered a gene as o A discrete unit of inheritance o A region of specific nucleotide sequence in a chromosome 0 A DNA sequence that codes for a specific polypeptide chain Overview Conduction the Genetic Orchestra 0 Prokaryotes and eukaryotes alter gene expression in response to their changing environment 0 In multicellular eukaryotes gene expression regulates development and is responsible for differences in cell types 0 RNA molecules play many roles in regulating gene expression in eukaryotes Bacteria often respond to environmental change by regulating transcription 0 Natural selection has favored bacteria that produce only the products needed by that cell 0 A cell can regulate the production of enzymes by feedback inhibition or by gene regulation 0 Gene expression in bacteria is controlled by the operon model Operons The Basic Concept 0 A cluster of functionally related genes can be under coordinated by a single onoff switch 0 The regulatory switch is a segment of DNA called an usually positioned within the promoter 0 An is the entire stretch of DNA that includes the operator the promoter and the genes that they control 0 The operon can be switched off by a protein The repressor prevents gene transcription by binding to the operator and blocking RNA polymerase The repressor is the product of a separate The repressor can be in an active or inactive form depending on the presence of other molecules A is a molecule that cooperates with a repressor protein to switch an operon off For example E coli can synthesize the amino acid tryptophan By default the trp operon is on the genes for tryptophan synthesis are transcribed When tryptophan is present it binds to the trp repressor protein which turns the operon off The repressor is active only in the presence of its corepressor tryptophan thus the trp operon is turned off repressed is tryptophan levels are high Repressible and Inducible Operons Two Types of Negative Gene Regulation A repressible operon is one that is usually on binding of a repressor to the operator shuts off transcription The trp operon is a repressible operon An inducible operon is one that is usually off a molecule called an inducer inactivates the repressor and turns on transcription The lac operon is an inducible operon and contains genes that code for enzymes used in the hydrolysis and metabolism of lactose By itself the lac repressor is active and switches the lac operon off A molecule called and inducer inactivates the repressor to turn the lac operon on Inducible enzymes usually function in catabolic pathways their synthesis is induced by a chemical signal Repressible enzymes usually function in anabolic pathways their synthesis is repressed by high levels of the end product Regulation of the trp and lac operons involves negative control of genes because operons are switched off by the active form of the repressor Positive Gene Regulation Some operons are also subject to positive control through stimulatory protein such as catabolite activator protein CAP and activator of transcription When glucose a preferred food source of E coli is scarce CAP is activated by binding with cyclic AMP Activated CAP attaches to the promoter of the lac operon and increases the affinity of RNA polymerase thus accelerating transcription When glucose levels increase CAP detaches from the lac operon and transcription returns to a normal rate CAP helps regulate other operons that encode enzymes used in catabolic pathways Regulation of Chromatin Structure 0 Genes Within highly packed heterochromatic are usually not expressed 0 Chemical modifications to histones and DNA of chromatic in uence both chromatic structure and gene expression Histone Modifications o In acetyl groups are attached to positively charged lysines in histone tains 0 This process loosens chromatic structure thereby promoting the initiation of transcription 0 The addition of methyl groups methylation can condense chromatic the addition of phosphate groups phosphorylation next to a methylated amino acid can loosen chromatic DNA Methylation 0 DNA methylation the addition of methyl groups to certain bases in DNA is associated With reduced transcription in some species 0 DNA methylation can cause longterm inactivation of genes in cellular differentiation 0 In methylation regulates expression of either the maternal or paternal alleles of certain genes at the start of development Mechanisms of PostTranscriptional Regulation 0 Transcription alone does not account for gene expression 0 Regulatory mechanisms can operate at various stages after transcription 0 Such mechanisms allow a cell to finetune gene expression rapidly in response to environmental changes RNA Processing 0 In different mRNA molecules are produced from the same primary transcript depending on which RNA segments are treated as exons and Which as introns mRNA Degredation o The life span of mRNA molecules in cytoplasm is a key to determining protein synthesis 0 Eukaryotic mRNA is more long lived than prokaryotic mRNA 0 The mRNA life span is determined in part by sequences in the leader and trainer regions Protein Processing and Degradation 0 After translation various types of protein processing including cleavage and the addition of chemical groups are subject to control 0 are giant protein complexes that bind protein molecules and degrade them Noncoding RNAs plav multiple roles in controlling gene expression 0 Only a small fraction of DNA codes for proteins rRNA and tRNA 0 A significant amount of the genome may be transcribed into noncoding RNAs 0 Noncoding RNAs regulate gene expression at two points mRNA translation and chromatic configuration Effects on mRNAs by MicroRNAs and Small Interfering RNAs o are small singlestranded RNA molecules that can bind to mRNA 0 These can degrade mRNA or block its translation 0 The phenomenon of inhibition of gene expression by RNA molecules is called RNAi is causes by 0 siRNAs and miRNAs are similar but form from different RNA precursors
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