Gene Activity

DNA, RNA, & Protein

3

OutlineOutline

Function of GenesFunction of Genes

One Gene-One Enzyme HypothesisOne Gene-One Enzyme Hypothesis

Genetic CodeGenetic Code

TranscriptionTranscription

Processing Messenger RNAProcessing Messenger RNA

TranslationTranslation

Transfer RNATransfer RNA

Ribosomal RNARibosomal RNA

Chain of events:

DNA→ RNA→ Synthesis of ProteinOccurs in ALL living things – from bacteria to humans

5

Function of GenesFunction of Genes

Genes Specify EnzymesGenes Specify EnzymesBeadle and Tatum:Beadle and Tatum:

­ Experiments on fungus Neurospora crassaExperiments on fungus Neurospora crassa

­ Proposed that each gene specifies the Proposed that each gene specifies the synthesis of one enzymesynthesis of one enzyme

­ One-gene-one-enzyme hypothesisOne-gene-one-enzyme hypothesis

Genes Specify a PolypeptideGenes Specify a PolypeptideA gene is a segment of DNA that specifies the A gene is a segment of DNA that specifies the sequence of amino acids in a polypeptide sequence of amino acids in a polypeptide

Suggests that genetic mutations cause Suggests that genetic mutations cause changes in the primary structure of a proteinchanges in the primary structure of a protein

6Protein Synthesis:Protein Synthesis:From DNA to RNA to ProteinFrom DNA to RNA to Protein

The mechanism of gene expressionThe mechanism of gene expression

DNA in genes specify information, but DNA in genes specify information, but information is not structure and functioninformation is not structure and function

Genetic info is expressed into structure & Genetic info is expressed into structure & function through protein synthesisfunction through protein synthesis

The expression of genetic info into structure The expression of genetic info into structure & function:& function:

DNA in gene controls the sequence of DNA in gene controls the sequence of nucleotides in an RNA moleculenucleotides in an RNA molecule

RNA controls the primary structure of a RNA controls the primary structure of a proteinprotein

7Sickle-Cell Disease in Humans

RNA – Ribonucleic AcidSingle stranded helixContains ribose sugar (not deoxyribose)Uracil replaces thymine3 types of RNA

1. mRNA (messenger RNA)

2. rRNA (ribosomal RNA)

3. tRNA (transfer RNA)

Messenger RNA (mRNA)Carries genetic information from DNATravel from nucleus to the ribosome Direct protein synthesis

Ribosomal RNA (rRNA)Helps join the mRNA codons to the tRNA

anticodons in the ribosomes. It produce enzymes needed to bond amino acids together during protein synthesis

Transfer RNA (tRNA)Transport amino acids to the ribosome

10Structure of RNA

11Overview of Gene Expression

TranscriptionStep 1: DNA molecule unzips. RNA polymerase

binds where mRNA will be synthesized.

Step 2: Free RNA nucleotides attach to DNA nucleotides by base pairing

Step 3: mRNA strand breaks from DNA

Step 4: mRNA leaves nucleus and goes to cytoplasm to the ribosomes

13

Transcription of mRNATranscription of mRNA

A single chromosomes consists of one very long molecule A single chromosomes consists of one very long molecule encoding hundreds or thousands of genesencoding hundreds or thousands of genes

The genetic information in a gene describes the amino acid The genetic information in a gene describes the amino acid sequence of a proteinsequence of a protein The information is in the base sequence of one side (the “sense” The information is in the base sequence of one side (the “sense”

strand) of the DNA moleculestrand) of the DNA molecule The gene is the functional equivalent of a “sentence”The gene is the functional equivalent of a “sentence”

The segment of DNA corresponding to a gene is unzipped to The segment of DNA corresponding to a gene is unzipped to expose the bases of the sense strandexpose the bases of the sense strand The genetic information in the gene is transcribed (rewritten) into The genetic information in the gene is transcribed (rewritten) into

an mRNA moleculean mRNA molecule The exposed bases in the DNA determine the sequence in which The exposed bases in the DNA determine the sequence in which

the RNA bases will be connected togetherthe RNA bases will be connected together RNA polymerase connects the loose RNA nucleotides togetherRNA polymerase connects the loose RNA nucleotides together

The completed transcript contains the information from the The completed transcript contains the information from the gene, but in a mirror image, or complementary formgene, but in a mirror image, or complementary form

Transcription- The process of transferring genetic information from DNA to RNA

forms one single-stranded RNA molecule

Codon- 3 base code in DNA or mRNA. Each base of a codon in DNA is transcribed in to the mRNA code.

16Transcription

The order of nitrogen bases in mRNA will determine the type and order of amino acids in a protein

The code is universal - the codons represent the same amino acids in all organisms

Three nucleotides code for one amino acid

Example – UUU codes for phenylalanine

18Steps in Gene Expression:Steps in Gene Expression:TranslationTranslation

tRNA molecules have two binding sitestRNA molecules have two binding sites One associates with the mRNA transcriptOne associates with the mRNA transcript

The other associates with a specific amino acidThe other associates with a specific amino acid

Each of the 20 amino acids in proteins associates with one or Each of the 20 amino acids in proteins associates with one or more of 64 species of tRNAmore of 64 species of tRNA

TranslationTranslation An mRNA transcript migrates to rough endoplasmic reticulumAn mRNA transcript migrates to rough endoplasmic reticulum

Associates with the rRNA of a ribosomeAssociates with the rRNA of a ribosome

The ribosome “reads” the information in the transcriptThe ribosome “reads” the information in the transcript

Ribosome directs various species of tRNA to bring in their Ribosome directs various species of tRNA to bring in their specific amino acid “fares”specific amino acid “fares”

tRNA specified is determined by the code being translated in tRNA specified is determined by the code being translated in the mRNA transcriptthe mRNA transcript

19

The Genetic CodeThe Genetic Code

The unit of a code consists of codons, each of which is a The unit of a code consists of codons, each of which is a unique arrangement of symbolsunique arrangement of symbols

Each of the 20 amino acids found in proteins is uniquely Each of the 20 amino acids found in proteins is uniquely specified by one or more codons specified by one or more codons The symbols used by the genetic code are the mRNA basesThe symbols used by the genetic code are the mRNA bases

­ Function as “letters” of the genetic alphabetFunction as “letters” of the genetic alphabet

­ Genetic alphabet has only four “letters” (U, A, C, G)Genetic alphabet has only four “letters” (U, A, C, G)

Codons in the genetic code are all three bases (symbols) longCodons in the genetic code are all three bases (symbols) long­ Function as “words” of genetic informationFunction as “words” of genetic information

­ Permutations:Permutations: There are 64 possible arrangements of four symbols taken three There are 64 possible arrangements of four symbols taken three

at a timeat a time Often referred to as tripletsOften referred to as triplets

­ Genetic language only has 64 “words”Genetic language only has 64 “words”

20

The Genetic CodeThe Genetic Code

Properties of the genetic code:Properties of the genetic code: UniversalUniversal

­ With few exceptions, all organisms use the code the With few exceptions, all organisms use the code the same waysame way

­ Encode the same 20 amino acids with the same 64 Encode the same 20 amino acids with the same 64 tripletstriplets

Degenerate (redundant)Degenerate (redundant)­ There are 64 codons available for 20 amino acidsThere are 64 codons available for 20 amino acids­ Most amino acids encoded by two or more codonsMost amino acids encoded by two or more codons

Unambiguous (codons are exclusive)Unambiguous (codons are exclusive)­ None of the codons code for two or more amino acidsNone of the codons code for two or more amino acids­ Each codon specifies only one of the 20 amino acidsEach codon specifies only one of the 20 amino acids

Contains start and stop signalsContains start and stop signals­ Punctuation codonsPunctuation codons­ Like the capital letter we use to signify the beginning of Like the capital letter we use to signify the beginning of

a sentence, and the period to signify the enda sentence, and the period to signify the end

•64 possible codons •20 different amino acids

Codons

Codons

Start CodonAUG

Stop CodonsUGAUAAUAG

23

Processing Messenger RNAProcessing Messenger RNA

Primary mRNA transcript is modified before it leaves Primary mRNA transcript is modified before it leaves the eukaryotic nucleusthe eukaryotic nucleus RNA splicing:RNA splicing:

­ Primary transcript consists of:Primary transcript consists of: Some segments that will not be expressed (introns)Some segments that will not be expressed (introns) Segments that will be expressed (exons)Segments that will be expressed (exons)

­ Performed by spliceosome complexes in nucleoplasmPerformed by spliceosome complexes in nucleoplasm Introns are excisedIntrons are excised Remaining exons are spliced back togetherRemaining exons are spliced back together

Modifications to ends of primary transcript:Modifications to ends of primary transcript:­ Cap of modified guanine on 5Cap of modified guanine on 5′′ end end­ Poly-A tail of 150+ adenines on 3Poly-A tail of 150+ adenines on 3′′ end end

Result is mature mRNA transcriptResult is mature mRNA transcript

24RNA Polymerase

25mRNA Processing in Eukaryotes

26

Functions of IntronsFunctions of Introns

As organismal complexity increases;As organismal complexity increases; Number of protein-coding genes does not keep paceNumber of protein-coding genes does not keep pace But the proportion of the genome that is introns But the proportion of the genome that is introns

increasesincreases Humans:Humans:

­ Genome has only about 25,000 coding genesGenome has only about 25,000 coding genes­ Up to 95% of this DNA genes is intronsUp to 95% of this DNA genes is introns

Possible functions of introns:Possible functions of introns: More bang for buckMore bang for buck

­ Exons might combine in various combinationsExons might combine in various combinations­ Would allow different mRNAs to result from one Would allow different mRNAs to result from one

segment of DNAsegment of DNA Introns might regulate gene expressionIntrons might regulate gene expression

Exciting new picture of the genome is emergingExciting new picture of the genome is emerging

RNA Editing (even better pic in your textbook p. 302)

28

tRNAtRNA

tRNA molecules come in 64 different kindstRNA molecules come in 64 different kinds

All very similar except thatAll very similar except that

One end bears a specific triplet (of the 64 One end bears a specific triplet (of the 64 possible) called the anticodonpossible) called the anticodon

Other end binds with a specific amino acid Other end binds with a specific amino acid typetype

tRNA synthetases attach correct amino acid to tRNA synthetases attach correct amino acid to the correct tRNA moleculethe correct tRNA molecule

All tRNA molecules with a specific anticodon All tRNA molecules with a specific anticodon will always bind with the same amino acidwill always bind with the same amino acid

29Structure of tRNA

30

RibosomesRibosomes

Ribosomal RNA (rRNA):Ribosomal RNA (rRNA):Produced from a DNA template in the Produced from a DNA template in the nucleolusnucleolus

Combined with proteins into large and small Combined with proteins into large and small ribosomal subunitsribosomal subunits

A completed ribosome has three binding sites A completed ribosome has three binding sites to facilitate pairing between tRNA and mRNAto facilitate pairing between tRNA and mRNAThe E (for exit) siteThe E (for exit) site

The P (for peptide) site, andThe P (for peptide) site, and

The A (for amino acid) siteThe A (for amino acid) site

31Ribosomal Structure and Function

32Steps in Translation:Steps in Translation:#1 - Initiation#1 - Initiation

Components necessary for initiation are:Components necessary for initiation are: Small ribosomal subunitSmall ribosomal subunit mRNA transcriptmRNA transcript Initiator tRNA, andInitiator tRNA, and Large ribosomal subunitLarge ribosomal subunit Initiation factors (special proteins that bring the Initiation factors (special proteins that bring the

above together)above together)

Initiator tRNA:Initiator tRNA: Always has the UAC anticodonAlways has the UAC anticodon Always carries the amino acid methionineAlways carries the amino acid methionine Capable of binding to the P siteCapable of binding to the P site

33Steps in Translation:Steps in Translation:#1 - Initiation#1 - Initiation

Small ribosomal subunit attaches to mRNA Small ribosomal subunit attaches to mRNA transcripttranscript

Beginning of transcript always has the START Beginning of transcript always has the START codon (AUG)codon (AUG)

Initiator tRNA (UAC) attaches to P siteInitiator tRNA (UAC) attaches to P site

Large ribosomal subunit joins the small Large ribosomal subunit joins the small subunitsubunit

34Steps in Translation:#1 - Initiation

35Steps in Translation:Steps in Translation:#2 - Elongation#2 - Elongation

““Elongation” refers to the growth in length of Elongation” refers to the growth in length of the polypeptidethe polypeptide

RNA molecules bring their amino acid fares to RNA molecules bring their amino acid fares to the ribosomethe ribosomeRibosome reads a codon in the mRNARibosome reads a codon in the mRNA

­ Allows only one type of tRNA to bring its amino Allows only one type of tRNA to bring its amino acidacid

­ Must have the anticodon complementary to the Must have the anticodon complementary to the mRNA codon being readmRNA codon being read

­ Joins the ribosome at it’s A siteJoins the ribosome at it’s A site

Methionine of initiator is connected to amino Methionine of initiator is connected to amino acid of 2acid of 2ndnd tRNA by peptide bond tRNA by peptide bond

36Steps in Translation:Steps in Translation:#2 – Elongation (cont.)#2 – Elongation (cont.)

Second tRNA moves to P site (translocation)Second tRNA moves to P site (translocation)

Spent initiator moves to E site and exitsSpent initiator moves to E site and exits

Ribosome reads the next codon in the mRNARibosome reads the next codon in the mRNA

Allows only one type of tRNA to bring its Allows only one type of tRNA to bring its amino acidamino acid

­ Must have the anticodon complementary to the Must have the anticodon complementary to the mRNA codon being readmRNA codon being read

­ Joins the ribosome at it’s A siteJoins the ribosome at it’s A site

Dipeptide on 2Dipeptide on 2ndnd amino acid is connected to amino acid is connected to amino acid of 3amino acid of 3ndnd tRNA by peptide bond tRNA by peptide bond

37Steps in Translation:#2 - Elongation

38Steps in Translation:Steps in Translation:#3 – Termination#3 – Termination

Previous tRNA moves to P sitePrevious tRNA moves to P siteSpent tRNA moves to E site and exitsSpent tRNA moves to E site and exitsRibosome reads the STOP codon at the end of Ribosome reads the STOP codon at the end of

the mRNAthe mRNAUAA, UAG, or UGAUAA, UAG, or UGADoes not code for an amino acidDoes not code for an amino acid

Polypeptide is released from last tRNA by Polypeptide is released from last tRNA by release factorrelease factor

Ribosome releases mRNA and dissociates Ribosome releases mRNA and dissociates into subunitsinto subunits

mRNA read by another ribosomemRNA read by another ribosome

39Steps in Translation:#3 - Termination

40Summary of Gene Expression

(Eukaryotes)

41

ReviewReview

Function of GenesFunction of Genes

One Gene-One Enzyme HypothesisOne Gene-One Enzyme Hypothesis

Genetic CodeGenetic Code

TranscriptionTranscription

Processing Messenger RNAProcessing Messenger RNA

TranslationTranslation

Transfer RNATransfer RNA

Ribosomal RNARibosomal RNA

Quick Molecular Genetic Terminology

Replication= DNA DNA Transcription= DNA RNA Translation= RNA Amino Acids Polypeptide= Chain of amino acids Adenine, cytosine, & guanine= DNA & RNA Thymine= DNA Only Uracil= RNA Only

SUMMARY

4444

Biology and Cells

All­living­organisms­consist­of­cells.­­Humans­have­trillions­of­cells.­­Yeast­-­one­cell.

Cells­are­of­many­different­types­(blood,­skin,­nerve),­but­all­arose­from­a­single­cell­(the­fertilized­egg)

Each*­cell­contains­a­complete­copy­of­the­genome­(the­program­for­making­the­organism),­encoded­in­DNA.

* there are a few exceptions

4545

DNA

DNA­molecules­are­long­double-stranded­chains;­4­types­of­bases­are­attached­to­the­backbone:­adenine­(A)­pairs­with­thymine­(T),­and­guanine­(G)­with­cytosine­(C).

A­gene­is­a­segment­of­DNA­that­specifies­how­to­make­a­protein.­

Proteins­are­large­molecules­are­essential­to­the­structure,­function,­and­regulation­of­the­body.­E.g.­are­hormones,­enzymes,­and­antibodies.­

E.g.­Human­DNA­has­about­30-35,000­genes;­­­­­­­­

­­­Rice­--­about­50-60,000,­but­shorter­genes.

4646

Exons and Introns: Data and Logic?

exons­are coding­DNA­(translated­into­a­protein),­which­are­only­about­2%­of­human­genome­­

introns­are­non-coding­DNA,­which­provide­structural­integrity­and­regulatory­(control)­functions­

exons­can­be­thought­of­program­data,­while­introns­provide­the­program­logic

Humans­have­much­more­control­structure­than­rice­

4747

Gene Expression

Cells­are­different­because­of­differential gene expression.

About­40%­of­human­genes­are­expressed­at­one­time.

Gene­is­expressed­by­transcribing­DNA­exons­into­single-stranded­mRNA

mRNA­is­later­translated­into­a­protein

4848

Molecular Biology Overview

Cell Nucleus

Chromosome

ProteinGraphics courtesy of the National Human Genome Research Institute

Gene (DNA)Gene (mRNA), single strand

Geneexpression