PowerPoint Presentation

CHAPTER 7GENETICS(Part 1)

Resilience expense 1CONTENT DNA ReplicationModels of DNA ReplicationDNA Replication Process

Genetic code and its features

Protein synthesis: Transcription and TranslationTranscription in prokaryotes and eukaryotesTranslation Initiation, Elongation and Termination steps.BIO095 (2013/2014)PART 1CONTENT Gene Regulation and expression

Mendelian geneticsMonohybrid inheritanceDihybrid inheritanceExtension of Mendelian Genetics

Population GeneticsGene populationHardy-Weinbergs PrincipleFactor that change the frequency in population BIO095 (2013/2014)(1) LEARNING OBJECTIVES Explain about the DNA replication process.

Summarize how DNA replicates (semiconservative) by citing the Meselson and Stahls experiment.

Outline the flow of genetic information process in cells, from DNA RNA Proteins via looking at the transcription and translation process.

BIO095 (2013/2014)(2) LEARNING OBJECTIVES Explain the concept of operon and gene regulation.

Describe the components of lac operon and their functions in E. coli.

Describe the mechanism of the operon in the absence and presence of lactose.

BIO095 (2013/2014)(3) LEARNING OBJECTIVES Define important terms in genetics.

Describe Mendel Principles of Segregation and Independent Assortment.

Explain population genetics, gene pool, allele frequencies and genetic equilibrium.

State and explain five assumptions of Hardy-Weinberg Law fro genetic equilibrium.

Calculate allele and genotype frequencies. BIO095 (2013/2014)

1. DNA REPLICATION:

DNA & RNA STRUCTUREBIO095 (2013/2014)Both DNA & RNA are macromolecules composed of smaller building blocks.

There are several levels of complexity needed to be considered:

Nucleotides forms the repeating structural unit of nucleic acids. Nucleotides are linked together in a linear manner to form a strand of DNA or RNA. polynucleotides 2 strands of DNA, interact with each other to form a double helix. The 3-dimensional structure of DNA results from the folding & bending of the double helix. Nucleic Acid StructureBIO095 (2013/2014)8Nucleotides 9

Single strand Double helix3-dimensional structureFig: Levels of nucleic acid structure9The nucleotide is the repeating structural unit of DNA & RNA.

3 components of a nucleotide:A phosphate groupA pentose sugarA nitrogenous base

There are 2 types of pentose sugars:Ribose found in RNADeoxyribose found in DNANucleotides: The Building Blocks of Nucleic AcidBIO095 (2013/2014)10Each nucleotide consists of 1 of the 5 types of nitrogenous bases.2 bases are derived from double-ringed purine,3 bases are derived from single-ringed pyrimidine.

Purine bases Double ringed carbon-nitrogen compoundAdenine (A)Guanine (G)

Pyrimidine bases Single ringed carbon-nitrogen compoundThymine (T)Cytosine (C)Uracil (U)Nucleotides: The Building Blocks of Nucleic AcidBIO095 (2013/2014)1112

Fig: The components of nucleotides12

DNA contains:deoxyribose as its sugarthe bases A, T, G, C

RNA contains:ribose as its sugar The bases A, U, G, C

Fig: The structure of nucleotides in (a) DNA and (b) RNARepeating unit of deoxyribonucleic acid (DNA)

Repeating unit of ribonucleic acid (RNA)13Nucleotides are covalently attached to each other in a linear manner to form a strand of DNA or RNA.

Structural features of DNA strand:Phosphodiester linkage the bond between a phosphate group on 1 nucleotide & the sugar molecule on the adjacent nucleotide.

The phosphates & the sugar molecules (sugar-phosphate backbone) form the backbone of DNA or RNA strand.

The backbone is negatively charged due to a negative charge on each phosphate. Nucleotides are linked together to form a strand

Fig: Nucleotide polymerBIO095 (2013/2014)Structural features of DNA strand:

4. The bases project from the backbone in the interior.

Phosphodiester linkage involves a phosphate attachment to the 5 carbon in 1 nucleotide & to the 3 carbon of adjacent nucleotide. The direction of the strand is 5 to 3 when going from top to bottom.

A strand of DNA or RNA contains a specific sequence of bases.

The nucleotides within the a strand are covalently attached to each other so the bases cannot shuffle around & become rearranged. .: the sequence of bases in a DNA strand will remain the same over time, except when mutations occur (rare cases).

The sequence of bases within DNA & RNA allows them to carry information.

14In a DNA double helix, 2 DNA strands are twisted together around a common axis.

There are 10 base pairs within a complete twist.

This double-stranded structure is stabilized by hydrogen bonding between the base pairs.

The AT/GC rule (a.k.a. Chargaffs rule) shows that,DNA from many organisms contains equal amounts of A & T, and equal amounts of G & C.

The Molecular Structure of the DNA Double HelixBIO095 (2013/2014)15There are:3 hydrogen bonds between G & C,2 hydrogen bonds between A & T.

For this reason, DNA sequences that have a high proportion of G & C tend to form more stable double-stranded structures.

The AT/GC rules implies that:base sequences within 2 DNA strands are complementary to each other.

Example:The sequence of 1st strand : 5-ATGGCGGATTT-3The sequence of the opposite strand : 3-TACCGCCTAAA-5The Molecular Structure of the DNA Double HelixBIO095 (2013/2014)The sequences are labeled with 5 (phosphate group) & 3 (-OH group) end.

These number designate the direction of the DNA backbones.

5 (phosphate group) & 3 (-OH group) end indicate the attachment to the carbon of the sugar. 16The polynucleotide has directionality from:5 (phosphate group) end to 3 (-OH group) end.

When going from the top of double-stranded DNA:The direction of the LEFT strand is: 5 3The direction of the RIGHT strand is: 3 5

Both strands are in antiparallel direction. i.e the 2 DNA strands are in opposite orientation.

An antiparallel structure was initially proposed in the models of Watson & Crick.The Molecular Structure of the DNA Double Helix

BIO095 (2013/2014)17

One complete turn 3.4 nm (34 )One nucleotide 0.34 nm (34 )2 nm (20 )5533

The double helix Watson & Crick model of DNAThe double helix model of a DNA molecule was discovered by Watson & Crick in 1953.

The diameter of DNA helix is 20 .The distance between base pairs is 3.4 .

A complete turn (360o) of the DNA helix consists of 10 base pairs with a distance of 34

The 2 strands of DNA form a right-handed double helix.

The bases in opposite strands hydrogen bond according to the AT/GC rule.

The 2 strands are antiparallel with regard to their 5 to 3 directionality.The double helix model of a DNA molecule was discovered by Watson & Crick in 1953.

The diameter of DNA helix is 20 .The distance between base pairs is 3.4 .

A complete turn (360o) of the DNA helix consists of 10 base pairs with a distance of 34

The 2 strands of DNA form a right-handed double helix.The bases in opposite strands hydrogen bond according to the AT/GC rule. The 2 strands are antiparallel with regard to their 5 to 3 directionality.

18The basic components of RNA are the same with DNA except for two major differences:The pyrimidine base uracil replace thymine Ribose replace deoxyribose

Adenine & Uracil form a base pair formed by 2 hydrogen bonds.

RNA exist as a single strand

RNA is important in the production of proteins in living organisms.

RNA STRUCTUREBIO095 (2013/2014)19RNA STRUCTURE

BIO095 (2013/2014)204 types of RNA:Ribosomal RNA (rRNA)Messenger RNA (mRNA)Transfer RNA (tRNA)Small nuclear RNA (snRNA)

All 4 types of RNA are made based onthe genetic information from the different parts of the DNA inside the nucleus with the help of specific enzymes.

RNA STRUCTUREBIO095 (2013/2014)21mRNA A linear sequence of mRNA that copy the information contained in DNA (genes) to be translated in the translation process to make protein.travels to the ribosomes (protein making factories)

rRNAis a component of the ribosomes, the protein synthetic factories in the cell

tRNAbrings amino acids (raw materials) to the protein factories (ribosome) that makes different proteins.

Different tRNA molecules for each of the different amino acids.

snRNA is the name used to refer to a number of small RNA molecules found in the nucleus.

RNA STRUCTUREBIO095 (2013/2014)22

BIO095 (2013/2014)23

ribosomePolypeptide chainrRNA + proteinLarge subunitsmall subunitTranslation of proteins involves a mRNA, tRNA, ribosome, and energy.BIO095 (2013/2014)241. DNA REPLICATION: Models of DNA Replication BIO095 (2013/2014)DNA replicationis the process of copying a double-stranded DNA molecule.

In a cell, DNA replication must happen before cell division.

Prokaryotesreplicate their DNA throughout the interval between cell divisions.

In eukaryotes, timings are highly regulated and this occurs during the S phase of the cell cycle, preceding mitosis or meiosis I.BIO095 (2013/2014)DNA Replication 26When a cell copies a DNA molecule,each strand serves as a template for ordering nucleotides into a new complementary strand.

Nucleotides line up along the template strand,according to the base-paring rules &are lined to form the new strands.

Each of the 2 daughter molecules have one old strand derived from the parent molecule; &one newly synthesized strand.

This is called the SEMICONSERVATIVE MODEL.

BIO095 (2013/2014)DNA Replication 27BIO095 (2013/2014)DNA Replication: SEMICONSERVATIVE MODEL

The parent molecule has 2 complementary strands of DNA.Each base is paired by hydrogen bonding with its specific partner, A with T & G with C. The first step in replication is separation of the 2 DNA strands.Each parental strand can serves as a template that determines the order of nucleotides along a new, complementary strand.

The complementary nucleotides line up & are connected to form the sugar-phosphate backbones of the new strands.Each daughter DNA molecule consists of 1 parental strand (dark blue) & 1 new strand (light blue)

D

28There are 3 alternatives models of DNA replication. Conservative modelSemiconservative modelDispersive model

In 1950s, Matthew Meselson & Franklin Stahl,Carry out experiment that supported the semiconservative model of DNA replication.

DNA Replication BIO095 (2013/2014)29

Conservative model The 2 parental strands reassociate after acting as templates for new strands,Thus restoring the parental double helix. Semiconservative model The 2 parental strands separate,& each functions as a template for the synthesis of new, complementary strand. Dispersive model Each strand of both daughter molecules contains a mixture of old & newly synthesized DNA. 30(1.) DNA REPLICATION: DNA Replication Process BIO095 (2013/2014)The replication of DNA molecule begins at origin of replication,i.e. short stretches of DNA having specific sequence of nucleotides.

Enzymes that initiate DNA replication, recognize this sequence & attach to the DNA separating the 2 strands & opening up a replication bubble.

BIO095 (2013/2014)DNA Replication: GETTING STARTED

32Replication of DNA proceeds in both directions,until the entire molecule is copied.

A eukaryotic chromosome,have hundreds to few thousands replication of origins.

Multiple replication bubble form & eventually fuse,thus speeding up the copying of very long DNA molecules.

BIO095 (2013/2014)DNA Replication: GETTING STARTED

The E. coli chromosome (n many other bacterial chromosomes) is circular & has a single origin.

33The E. coli chromosome (& many other bacterial chromosomes) is circular & has a single origin. Bacterial DNA replication is bidirectional since the chromosome is circular. It begins from a central origin and proceeds around the chromosome.

BIO095 (2013/2014)DNA Replication: GETTING STARTED

http://academic.pgcc.edu/~kroberts/Lecture/Chapter%207/replication.html

Bacterial DNA replication is bidirectional since the chromosome is circular. It begins from a central origin and proceeds around the chromosome until the two polymerase enzymes meet. The torsion placed on the separated strands by the untwisting activity of helicase is relaxed by the enzyme topoisomerase by cutting the twisting sections and re-joining them opposite to the direction of the supercoil.34At each end of replication bubble, is a replication fork.Replication fork a Y-shaped region where the parental strands of DNA are being unwound.

Several kinds of proteins participate in the unwinding:HelicasesSingle-strand binding proteinsTopoisomerasePrimase BIO095 (2013/2014)DNA Replication: GETTING STARTED 35HelicaseEnzymes that untwist the double helix at the replication forks,separating the 2 parental strands making them available as template strands.

Single-strand binding proteinsBind to unpaired DNA strands (just after the parental strand separation), stabilizing them.

TopoisomeraseHelps relieve the strain ahead of the replication fork due to the untwisting of double helix, by breaking, swiveling & rejoining DNA strands.

BIO095 (2013/2014)DNA Replication: GETTING STARTED TopoisomeraseHelps relieve the strain due to the untwisting of double helix, that causes tighter twisting & strain ahead of the replication fork. Topoisomerase relieve the strain by breaking, swiveling & rejoining DNA strands.

36BIO095 (2013/2014)DNA Replication: GETTING STARTED

Start to synthesize new DNA strandHelicase unwinds & separates the parental DNA strands

SSB Protein stabilize the unwound parental strands

Topoisomerase breaks, swivels, & rejoins the parental DNA ahead of the replication fork, relieving the strain caused by unwinding.

Primase synthesizes RNA primers using the parental DNA as a template. 37During DNA synthesis, the initial nucleotide chain produced is a short RNA chain called RNA primer. RNA primer is synthesized by primase.

PrimaseSynthesizes RNA primer, using the parental DNA as a template.Primase start an RNA chain from a single RNA nucleotide, adding RNA nucleotide 1 at a time.The completed primer usually 5 10 nucleotides long.

The new DNA strand will start from the 3 end of the RNA primer.

BIO095 (2013/2014)DNA Replication: GETTING STARTED 38DNA polymerases,catalyze the synthesis of new DNA, by adding nucleotides to a preexisting chains.

2 major DNA polymerases are:DNA polymerase IIIUses parental DNA as a template to synthesize new DNA strand by covalently adding nucleotides to the 3 end of a pre-existing DNA strand or RNA primer.

DNA polymerase IRemoves RNA nucleotides of primer from the 5end & replaces them with DNA nucleotides. (U T)

BIO095 (2013/2014)DNA Replication: Synthesizing a New DNA Strand

In E.coli there are several different DNA polymerases , but 2 play the major roles in DNA replication:DNA polymerase IDNA polymerase III

In eukaryotes more complicated, with at least 11 different DNA polymerases discovered so far but the general principles are the same. 39The 2 DNA strands in a double helix are antiparallel,i.e. they are oriented in opposite directions to each other.

The 2 new strands formed during DNA replication,must also be antiparallel to their template strands.

DNA polymerases can add nucleotides only to the 3 end of a primer or growing DNA strand,& never to the 5 end.

Thus, a new DNA strand can elongate only in the 5 3 direction. BIO095 (2013/2014)DNA Replication: ANTIPARALLEL ELONGATION40The synthesis of new DNA molecules during DNA replication involves:The synthesis of the leading strandsOccurs from the Origin of replication towards the replication fork.

The synthesis of lagging strands Occurs away form replication fork towards Origin of replication. BIO095 (2013/2014)DNA Replication: ANTIPARALLEL ELONGATION

41Along 1 template strand, starting at origin of replication (that has a 3 end )Primase synthesized a short RNA primer.

This enables the DNA polymerase III to synthesize a complementary strand continuously,by elongating the new DNA strand in the 5 3 direction towards replication fork.

DNA pol III settles in the replication fork on the template strand & continuously adds nucleotides to the new complementary strand as the fork progresses.

BIO095 (2013/2014)1. Synthesis of Leading Strand

42The DNA strand made by this mechanism is called the leading strand.

Only 1 primer is required for DNA pol III to synthesize the leading strand.

DNA pol I Then replaces the RNA primer with DNA nucleotides.

BIO095 (2013/2014)1. Synthesis of Leading Strand

43

After RNA primer is made,DNA pol III starts to synthesize the leading strand.Leading strand is elongated continuously in the 5 3 direction, as the fork progresses. 44To elongate the other new DNA strand in the 5 3 direction,DNA pol III elongates this DNA strand in the direction away from the replication fork. This DNA strand is called the lagging strand.

The lagging strand is synthesize discontinuously,as a series of segments.

These segments of the lagging strands are called Okazaki fragments.Each Okazaki fragment must be primed separately. This requires primase. BIO095 (2013/2014)2. Synthesis of Lagging StrandOkazaki fragments Named after the Japanese scientists who discovered them.

The fragments are about 1,000 to 2,000 nucleotides long in E. coli 100 to 200 nucleotides long in eukaryotes. 45DNA pol I then replaces the RNA nucleotides of the primers with DNA nucleotides.

DNA ligase joins the Okazaki fragments to form the lagging strand Into continuous DNA strand.

BIO095 (2013/2014)2. Synthesis of Lagging StrandOkazaki fragments Named after the Japanese scientists who discovered them.

The fragments are about 1,000 to 2,000 nucleotides long in E. coli 100 to 200 nucleotides long in eukaryotes. 46

1. Helicase unwinds parental double helix 2. SSB Proteins stabilise the unwound parental DNA 3. The leading strand is synthesized continually in the 5 3 direction by DNA pol IIIOverall Direction of replication4. The lagging strand is synthesized discontinuously (5 3)5. Primase synthesizes RNA primer, which is extended by DNA pol III to form an Okazaki fragment. 6. DNA pol I replaces the RNA primer with DNA. 7. DNA ligase joins the Okazaki fragments. 47http://www.wiley.com/college/pratt/0471393878/student/animations/dna_replication/index.html

BIO095 (2013/2014)2. Synthesis of Lagging StrandOkazaki fragments Named after the Japanese scientists who discovered them.

The fragments are about 1,000 to 2,000 nucleotides long in E. coli 100 to 200 nucleotides long in eukaryotes. 48(2.) GENETIC CODE AND ITS FEATURES

BIO095 (2013/2014)The information content in DNA (the genetic material), is in the form of specific nucleotides sequences along the DNA strands.

The DNA inherited by an organism, leads to specific traits by dictating the synthesis of proteins.

Proteins are the links between genotype & phenotype.

BIO095 (2013/2014)From Gene to Protein50Genes provide the instructions for making specific proteins. Examples:EnzymesStructural proteins e.g. keratin in hairHormone e.g. insulinGlobular protein e.g. hemoglobin

BUT a gene DOES NOT built a protein directly.

RNA - is the bridge between DNA & protein synthesis.

BIO095 (2013/2014)From Gene to Protein51To get from DNA (sequences of nucleotides) to proteins (sequences of amino acids),requires 2 major stages:

Transcription (DNA RNA)

Translation (RNA Protein)

BIO095 (2013/2014)From Gene to Protein52The synthesis of RNA under the direction of DNA

Both nucleic acids (DNA & RNA) use the same language i.e. Nucleotides

.: The information from DNA is transcribed or copied into RNA form.

DNA provides a template for assembling a sequence of RNA nucleotides. This produces messenger RNA (mRNA)

BIO095 (2013/2014)From Gene to Protein: TRANSCRIPTION53mRNA:is a transcript of the genes protein-building instructions (from the DNA)

carries a genetic message from the DNA to ribosomes (the protein-synthesizing machinery of the cell).

BIO095 (2013/2014)From Gene to Protein: TRANSCRIPTION54The synthesis of a polypeptide under the direction of mRNA.

The base sequence of an mRNA molecule is translated into the amino acid of a polypeptide.

Ribosomes,the site of translationfacilitate the orderly linking of amino acids into polypeptide chains.

BIO095 (2013/2014)From Gene to Protein: TRANSLATION55

Transcription & Translation in Eukaryotic cellThe inherited information flows from DNA RNA Protein. Transcription:Occurs in the nucleus.A gene provides the instruction for synthesizing mRNA.

RNA Processing:The original RNA transcript (primary transcript or pre-mRNA) is processed before leaving the nucleus as functional mRNA.

Translation:The information encoded in mRNA determines the sequence of amino acids to form a specific polypeptide. Occurs at ribosomes in the cytoplasm.

The flow of information from gene to protein is based on a triplet code.

Triplets of nucleotides bases can code for all the 20 amino acids.

An mRNA molecule:Is complementary to its DNA template (carries genes)because RNA bases are assemble on the template according to the base-pairing rules.

Codons:are the mRNA base triplets.Example: UGG is the codon for amino acid tryptophan (Trp)Can also be use for the complementary DNA base triplet. Example: DNA codon: ACC corresponding RNA codon: UGG

BIO095 (2013/2014)The Genetic Code: Nucleotide Triplets57

The Dictionary of The Genetic Code: Nucleotide Triplets & Its Amino AcidsCharacteristics of the genetic codeTriplet: Three nucleotides specify one amino acid61 codons correspond to amino acidsAUG codes for methionine and signals the start of transcription 3 stop codons signal the end of translation

58

The Triplet CodeFor each gene,1 DNA strand functions as a template for transcription. Follows the base-pairing rules.

During translation, the mRNA is read,as a sequence of base triplets codons,in the 5 3 direction

Each codon specifies an amino acid to be added to the growing polypeptide chain.

59(3.) Protein synthesis: Transcription and Translation TRANSCRIPTION

BIO095 (2013/2014)Overview of transcriptionInformation in DNA is transcribed (copied) to RNA molecule (mRNA molecule).

DNA act as a template,whereby a mRNA is transcribed complementary to the DNA template strand.

Like DNA replication, complementary fragments (mRNA) are generated in the 5' 3' direction.

Transcription leads to the translation of the genetic code (via the mRNA intermediate) into a functional peptide or protein.

RNA polymerase catalyzes the reactionBIO095 (2013/2014)TRANSCRIPTION OF RNA Transcription is the beginning of the process that leads to the translation of the genetic code (via the mRNA intermediate) into a functional peptide or protein.

61Stages of transcriptionINITIATIONRNA polymerase binds to a promoter, where the helix unwinds and transcription starts

ELONGATIONRNA nucleotides are added to the chain

TERMINATIONRNA polymerase reaches a terminator sequence and detaches from the templateBIO095 (2013/2014)The Synthesis & Processing of RNA:TRANSCRIPTION OF RNA 62PromoterThe DNA sequence where RNA polymerase attaches & initiates transcription.

Terminator The DNA sequence that signals the end of termination.

Transcription unitThe stretch of DNA that is transcribed into an RNA molecule.

BIO095 (2013/2014)The Synthesis & Processing of RNA:TRANSCRIPTION OF RNA In eukaryotes,RNA polymerase II used for mRNA synthesis

63The promoter of a gene,serves as a binding site for RNA polymerasedetermines where transcription startdetermines which of the 2 DNA strands is used as the template.

TATA boxA crucial promoter DNA sequence.

BIO095 (2013/2014)Stages of RNA Transcription : (1.) INITIATION

64RNA polymerase binds to the DNA at a specialized sequence called a promoter.

RNA polymerase unwinds DNA double helix and becomes single stranded.

RNA polymerase starts to transcribes the DNA with the addition of free RNA nucleotides in the 5 3 direction.(RNA polymerase add RNA nucleotides to the 3 end)

BIO095 (2013/2014)Stages of RNA Transcription : (1.) INITIATION *Adenine (A) in DNA complement with Uracil(U) in mRNA.Guanine (G) in DNA complement with Cytosine (C) in mRNA.Thymine (T) in DNA complement with Adenine (A) in mRNA.65RNA PolymeraseSeparates the 2 DNA strands apart.

add RNA nucleotides to the 3 end of the growing RNA polymeraccording to the base-pairing rules along the DNA template. .: RNA molecule elongates in 5 3 direction. BIO095 (2013/2014)

Stages of RNA Transcription : (1.) INITIATION 66RNA polymerase continues to moves along the DNA to unwind the double helix,

The enzyme,add nucleotides to the 3 end of the growing RNA molecule, as it moves along the double helix.

Then the DNA double helix reforms & the new RNA molecule peels away from its DNA template.

Transcription progresses at a rate of ~ 60 nucleotides per second in eukaryotes. BIO095 (2013/2014)Stages of RNA Transcription : (2.) ELONGATION 67Transcription proceeds until after the RNA polymerase transcribes a terminator sequence in the DNA.

Terminator sequence signals the end of the transcription unit.

The transcribed terminator (an RNA sequence) function as the actual termination signal.

The RNA transcript is released.

The RNA polymerase detaches from the DNA.

BIO095 (2013/2014)Stages of RNA Transcription : (3.) TERMINATION68

In prokaryotic cell, when the polymerase reaches at the end of the termination signal, transcription usually stops, & releases both the RNA & the DNA.

In eukaryotic cell, the polymerase continues for hundreds of nucleotides past the stop codon (the termination signal), i.e. an AAUAAA sequence in the pre-mRNA.

At a point ~ 10 35 nucleotides past the AAUAAA sequence,the pre-mRNA is cut free from the enzyme.

69Enzymes in the eukaryotic nucleus modify pre-mRNA, before the genetic message are transported to the cytoplasm.

RNA processing involves:Alteration of mRNA endsSplit genes & RNA splicing

BIO095 (2013/2014)MODIFYING RNA AFTER TRANSCRIPTION 70Each end of a pre-mRNA molecule is modified.

The 5 end of pre-mRNA The 5 end is capped off with 5 cap.5 cap is a modified guanosine triphosphate.

Function of 5 cap:Helps protect the mRNA from degradation by hydrolytic enzyme.After the mRNA reaches cytoplasm, there it functions as attach here sign for ribosomes.

BIO095 (2013/2014)1. ALTERATION OF MRNA ENDS71The 3 end of pre-mRNAAt the 3 end, an enzyme makes a Poly (A) tail.Poly (A) tail:Consists of ~ 50 250 adenine nucleotide.Inhibit degradation of RNA.Facilitate the export of mRNA from the nucleus.Helps ribosomes attach to it.

BIO095 (2013/2014)1. Alteration of mRNA ends72BIO095 (2013/2014)RNA processing: Addition of the 5 cap & Poly (A) tailThe leader, trailer & the poly (A) tail are not translated.

Trailer Leader Termination signal73DNA molecule contains non-coding sequences (the DNA regions that are not translated), that are interspaced between coding segment of the gene (DNA nucleotides sequence that codes for a polypeptide).

Thus the non-coding region can also be found between coding segment of the pre-mRNA.

RNA splicing:Is the removal of a large portion of the pre-mRNA moleculeBIO095 (2013/2014)2. SPLIT GENES & RNA SPLICING74Introns (intervening sequences):The non-coding segments of nucleic acid,Located between coding regions.

Exon:The coding segments,Are expressed by being translated into amino acid sequences.

The term intron & exon are used for both DNA & RNA.BIO095 (2013/2014)2. Split Genes & RNA Splicing75RNA polymerase transcribed both introns & exons from DNA to make pre-mRNA.

Introns are cut out from the molecule.

Exons are joined together to form an mRNA molecule with continuous coding sequence processed called RNA SPLICING. BIO095 (2013/2014)2. Split Genes & RNA Splicing76During RNA processing, the introns are excised, the exons are spliced together (RNA splicing) to form a continuous protein-coding message.

BIO095 (2013/2014)RNA Processing: RNA splicing

77

78(3.) Protein synthesis: Transcription and Translation TRANSLATION

BIO095 (2013/2014)Translation: The RNA directed synthesis of a polypeptide

The structure & function of transfer RNA

Ribosomes

Building a Polypeptide

BIO095 (2013/2014)THE SYNTHESIS OF PROTEINS80BIO095 (2013/2014)

Translation 81Transfer RNA (tRNA)Function to transfer amino acid from the cytoplasms amino acid pool to a ribosome.Has 2 ends:1 end carries a specific amino acid.Another end is an anticodon (a nucleotide triplet)ANTICODON base-pairs with a complementary codon on mRNA.

The ribosome adds each amino acid brought to it by tRNA to the growing end of a polypeptide chain.

BIO095 (2013/2014)STRUCTURE OF TRNA 82tRNA:are transcribed from DNA templates.

Made in the nucleus.

Travels to cytoplasm where translation occurs.

Consists of a single RNA strand (~80 nucleotides long)

Looks like a clove leaf.

The anticodon triplet is unique to each tRNA type.

Each tRNA molecule can be used repeatedlyBIO095 (2013/2014)Structure of tRNA

83An amino acid attachment site allows each tRNA to carry a specific amino acid

Anticodons are written in3 5, to align properly with mRNAs codon (5 3 direction).

Example:Anticodon : 3-AAG-5 mRNA codon : 5-UUC-3BIO095 (2013/2014)Structure of tRNA

84Made up of 2 subunits Large & small subunits.

Ribosomal subunits areconstructed of proteins & ribosomal RNA (rRNA) made in the nucleolus

The ribosomal subunits are exported via nuclear pores to the cytoplasm.

In the cytoplasm, the large & small subunits joins to form a functional ribosomes.

Most cells contains thousands of ribosomes. .: rRNA is the most abundant type of RNA.

BIO095 (2013/2014)RIBOSOMES 85BIO095 (2013/2014)Ribosomes structure

86Ribosome has a binding site for mRNA3 binding site for tRNA:The A site (Aminoacyl-tRNA site)Holds the tRNA carrying amino acid to be added to the chain.

The P site (Peptidyl-tRNA site)Holds the tRNA carrying the growing polypeptide chain.

The E site (Exit site) Discharged tRNA leaves ribosome from the E site.

BIO095 (2013/2014)RIBOSOMES 87BIO095 (2013/2014)Ribosomes structure

88Translation the synthesis of a polypeptide chain.

3 stages of translation:Initiation ElongationTermination

BIO095 (2013/2014)BUILDING A POLYPEPTIDE89This stage brings together:mRNAA tRNA bearing the 1st amino acid of the polypeptideThe 2 subunits of a ribosome.

BIO095 (2013/2014)1. INITIATION 90The process:A small ribosomal subunit binds:mRNA & a special initiator tRNA.

The initiation codon, AUG (on mRNA),Signals the start of translation.

The initiator tRNA (carries Methionine) attaches to the initiation codon.

BIO095 (2013/2014)1. INITIATION 91Small ribosomal subunit binds to mRNA molecule.

Anticodon of initiator tRNA, UAC base-pairs with the start codon, AUG (mRNA).

The tRNA carries methionine (Met).

BIO095 (2013/2014)1. INITIATION

92Then a large ribosomal subunit is attached to produce a translation initiation complex.Consists of:Small & large subunitmRNAInitiator tRNA

The initiator tRNA sits in the P site.

The vacant A site is ready for the next aminoacyl tRNA.

BIO095 (2013/2014)1. INITIATION Energy in the form of a GTP molecule is spend to form the initiation complex.

93BIO095 (2013/2014)1. INITIATION

94Amino acids are added 1 by 1 to the preceding amino acid.

3-step cycle:Codon recognitionPeptide bond formationTranslocation

BIO095 (2013/2014)2. ELONGATION95The mRNA codon in the A site forms H bonds with the anticodon of an incoming aminoacyl tRNA. i.e.: Anticodon of an incoming tRNA will complement with the codon on mRNA.

This tRNA enters into the A site. BIO095 (2013/2014)2. ELONGATION: (1) Codon RecognitionRequires the hydrolysis of 2 GTP molecules. 96An rRNA molecule of the large ribosomal subunit,Function as a ribozyme,Catalyzes the formation of a peptide bond that joins the:Polypeptide extending from the P site to the new amino acid in the A site.

(The polypeptide separates from the tRNA to which it was attached ( which is in the P site) & binds to the amino acid of the tRNA in the A site.)BIO095 (2013/2014)2. ELONGATION: (2) Peptide bond formation97The tRNA (with its attached polypeptide) in the A site,is translocated to the P site, taking the mRNA along with it. This brings the next codon to be translated into the A site.

The previous tRNA that was in the P site is moved to the E site & released from the ribosome.

The mRNA moves through the ribosome in 1 direction: 5 3 direction (on the mRNA)

The ribosome & the mRNA move relative to each other,Unidirectionally codon by codon. BIO095 (2013/2014)2. ELONGATION: (3) Translocation Hydrolysis of a GTP molecule provides Energy for this step.

98The Elongation Cycle

TranslocationCodon recognitionPeptide bond formation99The final stage of transcription.

Elongation continues until:A stop codon in the mRNA reaches the A site of ribosome.3 Stop codon: UAA, UAG & UGADo not code for amino acid.Act as signal to stop translation.

BIO095 (2013/2014)3. TERMINATION100Release factorA proteinBinds directly to the stop codon in the A site.Hydrolyzes the bond between the tRNA (in the P site) & the last amino acid of the completed polypeptide chain. The polypeptide is released.

The 2 ribosomal subunit & the other components of the translation assembly DISSOCIATE!!

BIO095 (2013/2014)3. TERMINATIONHydrolyzes the bond between the tRNA (in the P site) & the last amino acid of the completed polypeptide chain (that the tRNA holds).

101BIO095 (2013/2014)3. TERMINATION

102

103What does translation represent:DNA RNA or RNA protein?

Where does the information for producing a protein originate: DNA or RNA?

Which one has a linear sequence of codons:rRNA, mRNA, or tRNA?

Which one directly influences the phenotype: DNA, RNA, or protein? BIO095 (2013/2014)REVIEW104The end of Part 1!!