UNIT 3 Transcriptionand Protein Synthesis. Objectives Discuss the flow of information from DNA to RNA to Proteins Discuss the flow of information from.

UNIT 3UNIT 3

Transcription Transcription and and

Protein SynthesisProtein Synthesis

ObjectivesObjectives Discuss the flow of information from DNA to RNA Discuss the flow of information from DNA to RNA

to Proteinsto Proteins Explain transcriptionExplain transcription Differentiate introns and exonsDifferentiate introns and exons State functions of the noncoding regions of mRNAState functions of the noncoding regions of mRNA Describe post transcriptional modification of pre-Describe post transcriptional modification of pre-

RNARNA Distinguish between mRNA, tRNA and rRNADistinguish between mRNA, tRNA and rRNA Distinguish the roles of ribosomes, tRNA and Distinguish the roles of ribosomes, tRNA and

mRNA in protein synthesismRNA in protein synthesis Give detailed description of the process and steps Give detailed description of the process and steps

of translationof translation Describe application of SDS-Page gel Describe application of SDS-Page gel

electrophoresis to protein analysiselectrophoresis to protein analysis

DNA to RNA to ProteinsDNA to RNA to Proteins

DNA to RNA to Proteins DNA to RNA to Proteins Eukaryotes vs. ProkaryotesEukaryotes vs. Prokaryotes

In prokaryotes, transcription and In prokaryotes, transcription and translation are translation are coupledcoupled

translation begins while the mRNA translation begins while the mRNA is still being synthesized.is still being synthesized.

mRNA in ProkaryotesmRNA in Prokaryotes


In Eukaryotes, In Eukaryotes, transcription and transcription and translation are translation are spatiallyspatially and and temporallytemporally separated separated

transcription occurs in transcription occurs in the nucleus to produce the nucleus to produce a pre-mRNA molecule. a pre-mRNA molecule.

pre-mRNA is pre-mRNA is processed to produce processed to produce the mature mRNA, the mature mRNA, which exits the nucleus which exits the nucleus and is translated in the and is translated in the cytoplasm.cytoplasm.

mRNA in EukaryotesmRNA in Eukaryotes


TranscriptionTranscription occurs in four main stages: occurs in four main stages:

1) binding of RNA polymerase to DNA at a 1) binding of RNA polymerase to DNA at a promoterpromoter

2) initiation of transcription on the template 2) initiation of transcription on the template DNA strandDNA strand

3) elongation of the RNA chain3) elongation of the RNA chain

4) termination of transcription along with the 4) termination of transcription along with the release of RNA polymerase and the completed release of RNA polymerase and the completed RNA product from the DNA template. RNA product from the DNA template.

TranscriptionTranscription Binding of polymerases to the initiation site at the Binding of polymerases to the initiation site at the

promoter.promoter. Prokaryotic RNA polymerases can recognize the promoter Prokaryotic RNA polymerases can recognize the promoter

and bind to it directly, but eukaryotic RNA polymerases and bind to it directly, but eukaryotic RNA polymerases have to rely on other proteins called have to rely on other proteins called transcription transcription factorsfactors..

Unwinding of the DNA double helix by Unwinding of the DNA double helix by helicasehelicase. . Prokaryotic RNA polymerases have the helicase activity, Prokaryotic RNA polymerases have the helicase activity,

but eukaryotic RNA polymerases do not. but eukaryotic RNA polymerases do not. Unwinding of eukaryotic DNA is carried out by a specific Unwinding of eukaryotic DNA is carried out by a specific

transcription factor.transcription factor. Synthesis of RNA based on the sequence of the DNA Synthesis of RNA based on the sequence of the DNA

template strandtemplate strand.. RNA polymerases use nucleoside triphosphates (NTPs) to RNA polymerases use nucleoside triphosphates (NTPs) to

construct a RNA strand.construct a RNA strand. Termination of synthesis. Termination of synthesis.

Prokaryotes and eukaryotes use different signals to Prokaryotes and eukaryotes use different signals to terminate transcription. terminate transcription.

PromoterPromoter The DNA promoter region in prokaryotes is The DNA promoter region in prokaryotes is

a stretch of about 40 base pairs adjacent to a stretch of about 40 base pairs adjacent to and including the transcription start point. and including the transcription start point.

The essential features of the promoter are The essential features of the promoter are the start point (designated +1 and usually the start point (designated +1 and usually an A), the six-nucleotide -10 sequence, and an A), the six-nucleotide -10 sequence, and the six-nucleotide -35 sequence. the six-nucleotide -35 sequence.

The two key sequences are located The two key sequences are located approximately 10 nucleotides and 35 approximately 10 nucleotides and 35 nucleotides upstream from the start point. nucleotides upstream from the start point.

Prokaryotic promoterProkaryotic promoter

ElongationElongation During elongation, RNA polymerase During elongation, RNA polymerase

binds to about 30 base pairs of DNA binds to about 30 base pairs of DNA At any given time, about 18 base pairs of At any given time, about 18 base pairs of

DNA are unwound, and the most DNA are unwound, and the most recently synthesized RNA is still recently synthesized RNA is still hydrogen-bonded to the DNA, forming a hydrogen-bonded to the DNA, forming a short RNA-DNA hybrid (12 bp long, but short RNA-DNA hybrid (12 bp long, but it may be shorter) it may be shorter)

The total length of growing RNA bound The total length of growing RNA bound to the enzyme and/or DNA is about 25 to the enzyme and/or DNA is about 25 nucleotides. nucleotides.

TerminationTermination Requires a termination sequence that Requires a termination sequence that

triggers the end of transcription. triggers the end of transcription. Two classes exist:Two classes exist:

rho dependent rho dependent a short complementary GC-rich sequence a short complementary GC-rich sequence

(followed by several U residues) will form a (followed by several U residues) will form a "brake" that will help release the RNA "brake" that will help release the RNA polymerase from the template.polymerase from the template.

rho independent. rho independent. binding of rho to the mRNA releases it from binding of rho to the mRNA releases it from

the template. the template.

TerminationTermination

rho-dependent –rho-dependent –

requires a protein requires a protein called rho to bind called rho to bind to specific to specific sequence. sequence.


rho-independent - rho-independent - depends on a depends on a seqence in mRNA seqence in mRNA which forms a which forms a stem loop. stem loop.


Transcription in Transcription in EukaryotesEukaryotes

Although transcription in eukaryotes is similar to Although transcription in eukaryotes is similar to that in prokaryotes, the process is more complex. that in prokaryotes, the process is more complex.

Instead of one RNA polymerase, there are three :Instead of one RNA polymerase, there are three : RNA polymerase IRNA polymerase I (localized to the nucleolus) (localized to the nucleolus)

transcribes the rRNA precursor molecules.transcribes the rRNA precursor molecules. RNA polymerase IIRNA polymerase II produces most mRNAs and produces most mRNAs and

snRNAs.snRNAs. RNA polymerase IIIRNA polymerase III is responsible for the production is responsible for the production

of pre-tRNAs, 5SrRNA and other small RNAs. of pre-tRNAs, 5SrRNA and other small RNAs. The mitochondria and chloroplasts have their own RNA The mitochondria and chloroplasts have their own RNA

polymerases. polymerases.

Transcription complexTranscription complex

http://highered.mcgraw-hill.com/sitehttp://highered.mcgraw-hill.com/sites/0072437316/student_view0/chaptes/0072437316/student_view0/chapter18/animations.html#r18/animations.html#


Eukaryotic nuclear genes have three Eukaryotic nuclear genes have three

classes of promoters which are classes of promoters which are individual for the three types of RNA individual for the three types of RNA polymerases polymerases


Termination signals end the Termination signals end the transcription of RNA by RNA transcription of RNA by RNA polymerase I and RNA polymerase polymerase I and RNA polymerase III without the activity of hairpin III without the activity of hairpin structures as seen in prokaryotes. structures as seen in prokaryotes.

mRNA is cleaved 10 to 35 base-pairs mRNA is cleaved 10 to 35 base-pairs downstream of a AAUAAA sequence downstream of a AAUAAA sequence (which acts as a poly-A tail addition (which acts as a poly-A tail addition signal). signal).

RNA processing in RNA processing in EukaryotesEukaryotes

Ribosomal RNA processing Ribosomal RNA processing involves cleavage of multiple rRNAs involves cleavage of multiple rRNAs

from a common precursor, with from a common precursor, with nontranscribed spacers separate the nontranscribed spacers separate the units. units.

The transcription unit is transcribed by The transcription unit is transcribed by RNA polymerase I into a single long RNA polymerase I into a single long transcript (pre-rRNA) transcript (pre-rRNA)


Every tRNA gene is transcribed as a Every tRNA gene is transcribed as a precursor that must be processed precursor that must be processed into a mature tRNA molecule by:into a mature tRNA molecule by: removal of the leader sequence at the 5removal of the leader sequence at the 5΄́

endend replacement of two nucleotides at the 3 replacement of two nucleotides at the 3 ΄́

end by the sequence CCA (with which all end by the sequence CCA (with which all mature tRNA molecules terminate)mature tRNA molecules terminate)

chemical modification of certain bases chemical modification of certain bases excision of an intron. excision of an intron.


Transcription of eukaryotic pre-mRNAs Transcription of eukaryotic pre-mRNAs often proceeds beyond the 3often proceeds beyond the 3΄́ end of end of the mature mRNA. the mature mRNA.

An AAUAAA sequence located slightly An AAUAAA sequence located slightly upstream from the proper 3upstream from the proper 3΄́ end then end then signals that the RNA chain should be signals that the RNA chain should be cleaved about 10-35 nucleotides cleaved about 10-35 nucleotides downstream from the signal site, downstream from the signal site, followed by addition of a poly-A tail followed by addition of a poly-A tail catalyzed by poly(A) polymerase. catalyzed by poly(A) polymerase.


Messenger RNA in eukaryotesMessenger RNA in eukaryotes is first made as heterogeneous nuclear is first made as heterogeneous nuclear

mRNA / pre-mRNA then processed into mRNA / pre-mRNA then processed into mature mRNA through:mature mRNA through: the addition of a 5 prime cap - a guanosine the addition of a 5 prime cap - a guanosine

nucleotide methylated at the 7th positionnucleotide methylated at the 7th position addition of poly-A tails addition of poly-A tails the splicing out of introns. the splicing out of introns.

Introns and ExonsIntrons and Exons EukaryoticEukaryotic genes have interrupted coding genes have interrupted coding

sequences. sequences.

There are long sequences of bases within There are long sequences of bases within the protein-coding sequences of the gene the protein-coding sequences of the gene that do not code for amino acids in the final that do not code for amino acids in the final protein product. protein product.

The nocoding regions within the gene are The nocoding regions within the gene are called called intronsintrons (intervening sequences). (intervening sequences).

The The exonsexons (expressed sequences) are (expressed sequences) are part of the protein-coding sequence.part of the protein-coding sequence.

How introns are removedHow introns are removed

How introns are removedHow introns are removed

The intron loops out as The intron loops out as snRNPs (small nuclear snRNPs (small nuclear ribonucleoprotein ribonucleoprotein particles, complexes of particles, complexes of snRNAs and proteins) snRNAs and proteins) bind to form the bind to form the spliceosome.spliceosome.

The intron is excised, The intron is excised, and the exons are then and the exons are then spliced together.spliced together.

The resulting mature The resulting mature mRNA may then exit the mRNA may then exit the nucleus and be nucleus and be translated in the translated in the cytoplasm.cytoplasm.

Animation –Animation –How introns are removedHow introns are removed

http://highered.mcgraw-hill.com/sitehttp://highered.mcgraw-hill.com/sites/0072437316/student_view0/chaptes/0072437316/student_view0/chapter15/animations.html#r15/animations.html#

Introns and ExonsIntrons and Exons

A typical eukaryotic gene may have A typical eukaryotic gene may have multiple exons and introns and the multiple exons and introns and the numbers are quite variable.numbers are quite variable.

In many cases the lengths of the In many cases the lengths of the introns are much greater than those introns are much greater than those of the exon sequences. of the exon sequences.

For instance the ovalbumin gene For instance the ovalbumin gene contains about 7700 base pairs, contains about 7700 base pairs, 1859 of them in exons.1859 of them in exons.

Ovalbumin geneOvalbumin gene

Functions of IntronsFunctions of Introns

Evidence now exists that introns Evidence now exists that introns have many functions, including for have many functions, including for regulation and structural purposes, regulation and structural purposes, and that many of the roles now and that many of the roles now hypothesized for introns are hypothesized for introns are plausible but need further plausible but need further elucidation. elucidation.

Please read at least three of Please read at least three of the following articles for the following articles for

tutorialtutorial

http://www.sciencedaily.com/releases/2009/05/090528203730.htm









Roles of ribosomes, Roles of ribosomes, mRNA and tRNA in mRNA and tRNA in Protein SynthesisProtein Synthesis

Protein SynthesisProtein Synthesis

http://highered.mcgraw-hill.com/http://highered.mcgraw-hill.com/sites/0072437316/student_view0/sites/0072437316/student_view0/chapter15/animations.html#chapter15/animations.html#

References/ sources of References/ sources of imagesimages

http://evolution.berkeley.edu/evolibrary/article/0_0_0/mutations_03http://evolution.berkeley.edu/evolibrary/article/0_0_0/mutations_03 usmlemd.wordpress.com/2007/07/14/dna-replication/usmlemd.wordpress.com/2007/07/14/dna-replication/ http://employees.csbsju.edu/hjakubowski/classes/ch331/dna/centraldogma.jpghttp://employees.csbsju.edu/hjakubowski/classes/ch331/dna/centraldogma.jpg http://www.usask.ca/biology/rank/demo/replication/cons.rep.gifhttp://www.usask.ca/biology/rank/demo/replication/cons.rep.gif http://click4biology.info/c4b/3/images/3.4/SEMICON.gifhttp://click4biology.info/c4b/3/images/3.4/SEMICON.gif http://www.bio.miami.edu/~cmallery/150/gene/sf12x16.jpghttp://www.bio.miami.edu/~cmallery/150/gene/sf12x16.jpg http://publications.nigms.nih.gov/findings/sept08/images/hunt_gene_big.jpghttp://publications.nigms.nih.gov/findings/sept08/images/hunt_gene_big.jpg http://ghr.nlm.nih.gov/handbook/illustrations/duplication.jpghttp://ghr.nlm.nih.gov/handbook/illustrations/duplication.jpg http://images.google.com.jm/imgres?imgurl=http://ghr.nlm.nih.gov/handbook/http://images.google.com.jm/imgres?imgurl=http://ghr.nlm.nih.gov/handbook/

illustrations/duplication.jpg&imgrefurl=http://ghr.nlm.nih.gov/handbook/illustrations/illustrations/duplication.jpg&imgrefurl=http://ghr.nlm.nih.gov/handbook/illustrations/duplication&usg=__BgKRLXXos-duplication&usg=__BgKRLXXos-xRaUqN5EyP7qchszc=&h=400&w=370&sz=38&hl=en&start=2&tbnid=ZfARmmvAKGxRaUqN5EyP7qchszc=&h=400&w=370&sz=38&hl=en&start=2&tbnid=ZfARmmvAKG02xM:&tbnh=124&tbnw=115&prev=/images%3Fq%3Dduplication%2Bmutation%26gbv02xM:&tbnh=124&tbnw=115&prev=/images%3Fq%3Dduplication%2Bmutation%26gbv%3D2%26hl%3Den%26client%3Dfirefox-a%26rls%3Dorg.mozilla:en-US:official%26sa%3D2%26hl%3Den%26client%3Dfirefox-a%26rls%3Dorg.mozilla:en-US:official%26sa%3DG%3DG

http://images.google.com.jm/imgres?imgurl=http://www.phschool.com/science/http://images.google.com.jm/imgres?imgurl=http://www.phschool.com/science/biology_place/biocoach/images/transcription/euovrvw.gif&imgrefurl=http://biology_place/biocoach/images/transcription/euovrvw.gif&imgrefurl=http://www.phschool.com/science/biology_place/biocoach/transcription/tctlpreu.html&usg=__-www.phschool.com/science/biology_place/biocoach/transcription/tctlpreu.html&usg=__-hoX0ehn3x9zc2nUeciZ8gLYLqQ=&h=288&w=261&sz=20&hl=en&start=10&um=1&thoX0ehn3x9zc2nUeciZ8gLYLqQ=&h=288&w=261&sz=20&hl=en&start=10&um=1&tbnid=X11pPXboEKhRIM:&tbnh=115&tbnw=104&prev=/images%3Fq%3Dtranscriptionbnid=X11pPXboEKhRIM:&tbnh=115&tbnw=104&prev=/images%3Fq%3Dtranscription%26ndsp%3D18%26hl%3Den%26sa%3DG%26um%3D1%26ndsp%3D18%26hl%3Den%26sa%3DG%26um%3D1

www.asa3.org/ASA/PSCF/2001/PSCF9-01Bergman.htmlwww.asa3.org/ASA/PSCF/2001/PSCF9-01Bergman.html http://www.mun.ca/biology/desmid/brian/BIOL2060/BIOL2060-21/CB21.htmlhttp://www.mun.ca/biology/desmid/brian/BIOL2060/BIOL2060-21/CB21.html http://www.mun.ca/biology/desmid/brian/BIOL2060/BIOL2060-21/2101.jpghttp://www.mun.ca/biology/desmid/brian/BIOL2060/BIOL2060-21/2101.jpg

UNIT 3 Transcriptionand Protein Synthesis. Objectives Discuss the flow of information from DNA to RNA to Proteins Discuss the flow of information from.

Documents

synthesis of rna

rna strand

eukaryotes dna

synthesized rna

rna chain

proteins dna

prokaryotes dna

short rnadna hybrid