Presentation on theme: "Genes: Structure Replication and Expression"— Presentation transcript:
1 Genes: Structure Replication and Expression Chapter 12Genes: Structure Replication and Expression
2 Replication:During mitotic division information is duplicated by DNA replication and is passed on to next generationdaughter cells has exavcyt replica of the parent DNA
3 Role of DNA in Protein synthesis DNA and protein synthesis involves:Transcription- yields a ribonucleic acid (RNA) copy of specific genesTranslation- uses information in messenger RNA (mRNA) to synthesize a polypeptide.Protein synthesis is assisted by RNA (tRNA) and ribosomal RNA (rRNA)
5 Nucleic Acid Structure Deoxyribonucleic Acid (DNA) polymer of nucleotidescontains the bases adenine, guanine, cytosine and thyminesugar is deoxyribosemolecule is usually double stranded
6 DNA is a double-stranded molecule twisted into a helix (think of a spiral staircase). Each spiraling strand, comprised of a sugar-phosphate backbone and attached bases, is connected to a complementary strand by non-covalent hydrogen bonding between paired bases.The bases are adenine (A), thymine (T), cytosine (C) and guanine (G). A and T are connected by two hydrogen bonds. G and C are connected by three hydrogen bonds.
9 DNA Structure – Two Complementary Strands base pairingAdenine (purine) and thymine (pyrimidine) pair by 2 hydrogen bondsGuanine (purine) and cytosine (pyrimidine) pair by 3 hydrogen bondsmajor and minor grooves form when the 2 strands twist around each other
10 Nucleic Acid Structure Ribonucleic Acid (RNA) polymer of nucleotidescontains the bases adenine, guanine, cytosine and uracilsugar is ribosemost RNA molecules are single stranded
11 RNA Structurethree different types which differ from each other in function and in structuremessenger RNA (mRNA)ribosomal RNA (rRNA)transfer RNA(tRNA)
12 The Organization of DNA in Cells In most bacteria DNA is a circular, double helixfurther twisting results in supercoiled DNAin bacteria the DNA is associated with basic proteinshelp organize the DNA into a coiled chromatin like structure
14 DNA Replicationcomplex process involving numerous proteins which help ensure accuracythe 2 strands separate, each serving as a template for synthesis of a complementary strandsynthesis is semi-conservative; each daughter cell obtains one old and one new strand
15 DNA Replicationbidirectional from a single origin of replication
16 DNA replication (arrows) occurs in both directions from the origin of replication in the circular DNA found in most bacteria.
17 Rolling Circle Replication some small circular genomes (e.g., viruses and plasmids)replicated by rolling-circle replicationAnimation illustrating DNA replication by complementary base pairingA single-stranded tail, often composed of more than one genome copy, is generated and can be converted to the double-stranded form by synthesis of a complementary strand. The “free end” of the rolling circle strand is probably bound to the primosome.
19 Gene Structure Gene reading frame the basic unit of genetic informationalso defined as the nucleic acid sequence that codes for a polypeptide, tRNA or rRNAlinear sequence of nucleotidescodons are found in mRNA and code for single amino acidsreading frameorganization of codons such that they can be read to give rise to a gene product
21 Genes that Code for Proteins template strand directs RNA synthesispromoter is located at the start of the geneis the recognition/binding site for RNA polymerasefunctions to orient polymeraseleader sequence is transcribed into mRNA but is not translated into amino acidsShine-Delgarno sequence important for initiation of translation
22 Genes that Code for Proteins The Coding Region:begins with the DNA sequence from 3´-TAC-5´produces codon AUG which codes for N-formylmethionine, a modified amino acid used to initiate protein synthesis in bacteria ( check fig.)coding region ends with a stop codon, immediately followed by the trailer sequence which contains a terminator sequence used to stop transcription
24 Genes That Code for tRNA and rRNA tRNA/rRNA genes have promoter (recognition/binding site for RNA polymerase), leader (is transcribed into mRNA), coding region, spacer and trailer regions (contains a terminator sequence used to stop transcription)during maturation process.leader, spacer, and trailer removed during maturation processFigure 12.19a:
25 rRNA genes have promoter, leader, coding, spacer, and trailer regions spacer and trailer regions may encodetRNA moleculesFigure 12.19b:
29 Transcription RNA is synthesized under the direction of DNA RNA produced has complementary sequence to the template DNAthree types of RNA are producedmRNA carries the message for protein synthesistRNA carries amino acids during protein synthesisrRNA molecules are components of ribosomes
30 Transcription in Bacteria… Definitions to understand protein synthesis:in most bacterial RNA polymerases:Holoenzyme can begin transcription> What is Holoenzyme??*the core enzyme is composed of 5 chains and catalyzes RNA synthesisthe sigma factor has no catalytic activity but helps the core enzyme recognize the start of genes*holoenzyme = core enzyme + sigma factoronly the holoenzyme can begin transcription
31 Transcription in Bacteria…. Transcription in Bacteria is catalyzed by a single RNA polymerase.a reaction similar to that catalyzed by DNA polymerase for DNA syntehsis.ATP,GTP,CTP and UTP are used to produce a complementary RNA copy of the template DNA sequence
34 Transcription Initiation Promotersite where RNA polymerase binds to initiate transcription & is not transcribed
35 Transcription Elongation after binding, RNA polymerase unwinds the DNAtranscription bubble producedmoves with the polymerase as it transcribes mRNA from template strandwithin the bubble a temporary RNA:DNA hybrid is formed
36 Coupled Transcription and Translation in Prokaryotes
38 The Genetic CodemRNA sequence is translated into amino acid sequence of polypeptide chain (process = translation).an understanding of the genetic code is necessary before translation is studied.
39 Organization of the Code code degeneracyup to six different codons can code for a single amino acidsense codonsthe 61 codons that specify amino acidsstop (nonsense) codonsthe three codons used as translation termination signalsdo not encode amino acids
41 Translation Translation of mRNA into protein: synthesis of polypeptide is directed by sequence of nucleotides in mRNARibosome:70S ribosomes = 30S + 50S subunitsite of translationpolyribosome (polysome) – complex of mRNA with several ribosomes
42 Translation of mRNA into protein: Three phases:InitiationElongationTermination
43 During translation, the mRNA is "read" according to the genetic code which relates the DNA sequence to the amino acid sequence in proteinsEach group of three base pairs in mRNA constitutes a codon, and each codon specifies a particular amino acid (hence, it is a triplet code).The mRNA sequence is thus used as a template to assemble—in order—the chain of amino acids that form a protein.
44 Transfer RNA (tRNA) and Amino Acid Activation The tRNA molecules are adaptor molecules—they have one end that can read the triplet code in the mRNA through complementary base-pairing, and another end that attaches to a specific amino acidattachment of amino acid to tRNA is catalyzed by aminoacyl-tRNA synthetases
45 The translation of mRNA begins with the formation of a complex on the mRNA (Fig. below). First, three initiation factor proteins (known as IF1, IF2, and IF3) bind to the small subunit of the ribosome.This preinitiation complex and a methionine-carrying tRNA then bind to the mRNA, near the AUG start codon, forming the initiation complex.
47 Methionine (Met) is the first amino acid incorporated into any new protein, however, it is not always the first amino acid in translation of protein.In many proteins, methionine is removed after translation.
48 The large ribosomal subunit binds to this complex, which causes the release of IFs (initiation factors) once the initiation complex is formed on the mRNAThe large subunit of the ribosome has three sites at which tRNA molecules can bind:The A (amino acid) site is the location at which the aminoacyl-tRNA anticodon base pairs up with the mRNA codon, ensuring that correct amino acid is added to the growing polypeptide chain.
49 The P (polypeptide) site is the location at which the amino acid is transferred from its tRNA to the growing polypeptide chain.Finally, the E (exit) site is the location at which the "empty" tRNA sits before being released back into the cytoplasm to bind another amino acid and repeat the process.
50 The initiator methionine tRNA is the only aminoacyl-tRNA that can bind in the P site of the ribosome, and the A site is aligned with the second mRNA codon.The ribosome is thus ready to bind the second aminoacyl-tRNA at the A site, which will be joined to the initiator methionine by the first peptide bond.
51 Elongation of the Polypeptide Chain The next phase in translation is known as the elongation phase .Elongation cycle is the sequential addition of amino acids to growing polypeptide & consists of three phasesaminoacyl-tRNA bindingtranspeptidation reactionTranslocationThe above process need several Elongation factors ( EF)
52 ………ElongationFirst, the ribosome moves along the mRNA in the 5'-to-3'direction, which requires the elongation factor G, in a process called translocation
53 …..Elongation CycleThe tRNA which corresponds to the second codon can then bind to the A site, a step that requires elongation factors (in E. coli, these are called EF-Tu and EF-Ts) and GTP (guanosine triphosphate ) as an energy source for this acitivity.Upon binding of the tRNA-amino acid complex in the A site, GTP is cleaved to form guanosine diphosphate (GDP), then released along with EF-Tu to be recycled by EF-Ts for the next round.elongation cycle of protein synthesis
54 ……….ElongationIn the next step, peptide bonds between the now-adjacent first and second amino acids are formed through a peptidyl transferase activity.After the peptide bond is formed, the ribosome shifts, or translocates, again, thus causing the tRNA to occupy the E site.The tRNA is then released to the cytoplasm to pick up another amino acid.The A site is now empty and ready to receive the tRNA for the next codon.
55 ….ElongationThis process is repeated until all the codons in the mRNA have been read by tRNA molecules &the amino acids attached to the tRNAs have been linked together in the growing polypeptide chain in the appropriate order.At this point, translation must be terminated, and the nascent protein must be released from the mRNA and ribosome.
56 Final Phase in Elongation Cycle − Translocation Three simultaneous events:peptidyl-tRNA moves from A site to P siteribosome moves down one codonempty tRNA leaves P site
57 Termination of Translation/protein synthesis: Three termination codons ( Non-sense or stop codon) that are employed at the end of a protein-coding sequence in mRNA: UAA, UAG, and UGANo tRNAs recognize these codons.Instead, release factors (RFs) helps in recognition of stop codons.Release factors are protein which binds and facilitates release of the mRNA from the ribosome and subsequent dissociation of the ribosome.
58 Several ribosome can align on one mRNA strand and forms several polypeptide chains each with 20 or more amino acids.
59 Prokaryotic and Eukaryotic Translation The translation process is very similar in prokaryotes and eukaryotes.Although different elongation, initiation, and termination factors are used, the genetic code is generally identical.In bacteria, transcription and translation take place simultaneously, and mRNAs are relatively short-lived.
60 In eukaryotes, mRNAs have highly variable half-lives, are subject to modifications, and must exit the nucleus to be translated.