1 genes expression or genes and how they work: transcription, translation, & more chapter 15

1

Genes Expression or Genes and How They Work: Transcription, Translation, & More

Chapter 15

2

Overview: Central Dogma

• Central Dogma– DNA RNA Protein

• During polypeptide synthesis, ribosomal RNA (rRNA) is the site of polypeptide assembly.

– Transfer RNA (tRNA) transports and positions amino acids.

– Messenger RNA (mRNA) directs which amino acids are assembled into polypeptides.

3

The Central Dogma

RNA

Protein

DNAProposed by Francis Crick in 1958 to describe the flow of information in a cell.

Information stored in DNA is transferredresidue-by-residue to RNA which in turn transfers the information residue-by-residue to protein.

The Central Dogma was proposed by Crick to help scientists think about molecular biology. It has undergone numerous revisions in the past 45 years.

Central Dogma

4

Central Dogma of Gene Expression

5

Transcription Overview

Def: DNA sequence is transcribed into RNA sequence

– initiated when RNA polymerase binds to promoter binding site

moves along DNA strand and adds corresponding complementary RNA nucleotide

disengages at stop signal

6

Translation Overview

• Def: nucleotide sequence of mRNA transcript is translated into amino acid sequence in the polypeptide

rRNA recognizes and binds to start sequence

moves three nucleotides at a time disengages at stop signal

• Gene expression - collective of transcription and translation

7

Genetic Code

• How does the order of nucleotides in a DNA molecule encode the information that specifies the order of amino acids in a polypeptide?

• The answer came in 1961 through an experiment lead by Crick.

8

Genetic Code

• Crick and colleagues reasoned that there must be codons or block of info that coded for an amino acid

• They hypothesized that it was most likely 3 nucleotides

– Why 3?– 2 nucleotides did not have enough combinations

(42 is only 16 possible amino acids)– 3 nucleotides (43 is 64 which is enough to cover

the roughly 20 known amino acids)

9

Genetic Code

• Now known• Genetic code consists of a series of

information blocks called codons.– reading frame (triplet)

each codes for one amino acid highly redundant

10

Genetic Code

11

amino group

20 amino acids

carboxyl group

12

Genetic Code

• Could be punctuated or not– Punctuated code would have a something

in the code that separates codons– Non-punctuated code would not

• In the following example, O is not a base pair but the “punctuation.”

13

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

15_06.jpg

(Nonsense)

(Nonsense)

Hypothesis A :unpunctuated

Delete 1 base

Delete T WHYDIDTHEREDBATEATTHEFATRAT?

WHYODIDOTHEOREDOBATOEATOTHEOFATORAT?

WHY DID HER EDB ATE ATT HEF ATR AT?

(Sense)

Hypothesis A :unpunctuated

Delete 3 bases

Delete T,R,and AWHYDIDTHEREDBATEATTHEFATRAT?

WHY DID HEE DBT EAT THE FAT RAT?

(Nonsense)

Hypothesis B :punctuated

Delete T

WHY DID HEO EDO ATO ATO HEO ATO AT?O O R B E T F R

WHYODIDOTHEOREDOBATOEATOTHEOFATORAT?

(Nonsense)

Hypothesis B :punctuated

Delete T,R,and A

WHY DID HEO DOB OEA OTH OFA ORA?O O E T T E T T

14

Genetic Code

• Could be punctuated or not– Punctuated code would have a something

in the code that separates codons– Non-punctuated code would not

• In the following example, O is not a base pair but the “punctuation.”

• Crick concluded that it is not punctuated as mutations lead to a different sequence of amino acids, not nonsense.

15

Genetic Code

• Code is practically universal– ex: AGA codes for arginine in bacteria,

humans and all other organisms studied– great evidence that all life has a common

ancestor– Genes coded in one organism can be

transcribed in another SWEET biotechnology

16

Genetic Code

• Code is practically universal…but not quite• In 1979 mammalian mitochondria found to

have different “universal code”– In mitochondrial DNA, UGA is not a stop

codon as it is in “universal code”– Other codons are different– Chloroplasts and ciliates (protists) have

minor differences as well• It is thought that the changes to

mitochondria and chloroplasts happen after their endosymbiotic existence

17

More on RNA

• Central Dogma shows how information is passed from DNA RNA Protein

• RNA’s structure is different from DNA

– single stranded not Double Stranded

– uracil not Thymine– ribose not

deoxyribose

18

• RNA like DNA, is a nucleic acid. RNA structure differs from DNA structure.

• First, RNA is single stranded—it looks like one-half of a zipper—whereas DNA is double stranded.

RNA StructureRNA Structure

19

• The sugar in RNA is ribose; DNA’s sugar is deoxyribose.

Ribose


20

• Both DNA and RNA contain four nitrogenous bases, but rather than thymine, RNA contains a similar base called uracil (U).

• Uracil forms a base pair with adenine in RNA, just as thymine does in DNA.

Uracil

Hydrogen bonds Adenine


21

Transcription

• RNA polymerase– only one of two DNA strands (template or

antisense strand) is transcribed– non-transcribed strand is termed coding

strand or sense strand– In both bacteria and eukaryotes, the

polymerase adds ribonucleotides to the growing 3’ end of an RNA chain.

synthesis proceeds in 5’3’ direction

22

Transcription Bubble

23

Transcription

• Promoter– Transcription starts at RNA polymerase

binding sites called promoters on DNA template strand.

• Initiation– Other eukaryotic factors bind, assembling

a transcription complex. RNA polymerase begins to unwind DNA

helix.

24

Transcription

• Elongation– Transcription bubble moves down DNA at

constant rate leaving growing RNA strands protruding from the bubble.

• Termination– Stop sequences at the end of the gene

cause phosphodiester bond formation to cease, transcription bubble to dissociate, and RNA polymerase to release DNA.

25

Transcription Complex

26

Eukaryotic Transcription

• Eukaryotic transcription differs from prokaryotic transcription:

– three RNA polymerase enzymes– initiation complex forms at promoter– RNAs are modified after transcription

27

Translation: From mRNA to Protein

• The process of converting the information in a sequence of nitrogenous bases in mRNA into a sequence of amino acids in protein is known as translation.

• Translation takes place at the ribosomes in the cytoplasm.

28


• The role of transfer RNA • For proteins to be built, the 20

different amino acids dissolved in the cytoplasm must be brought to the ribosomes.

• This is the role of transfer RNA (tRNA)

• Each tRNA molecule attaches to only one type of amino acid.

Amino acid

Chain of RNA nucleotides

Transfer RNA molecule

Anticondon

29

Ribosome

mRNA codon


30

Translation: From mRNA to Protein• The first codon on mRNA is AUG, which codes for the

amino acid methionine• AUG signals the start of protein synthesis.• When this signal is given, the ribosome slides along the

mRNA to the next codon.

tRNA anticodon

Methionine

31

• A new tRNA molecule carrying an amino acid pairs with the second mRNA codon.

Alanine


32

• The amino acids are joined when a peptide bond is formed between them.

AlanineMethionine

Peptide bond


33


• A chain of amino acids is formed until the stop codon is reached on the mRNA strand.

Stop codon

34

Translation (in more detail)

• Begins when initial portion of mRNA molecule binds to rRNA in a ribosome

– tRNA molecule with complimentary anticodon binds to exposed codon on mRNA

some tRNA molecules recognize more than one codon

35


• Activating enzymes– tRNA molecules attach to specific amino

acids through the action of activating enzymes (aminoacyl-tRNA syntheases).

must correspond to specific anticodon sequences on a tRNA molecule as well as particular amino acids

36


• Start and stop signals– start signal coded by AUG codon– stop signal coded by one of three

nonsense codons: UAA - UAG – UGA– What do you think “nonsense codons”

means here? • Initiation

– Polypeptide synthesis begins with the formation of an initiation complex.

initiation factors

37

Formation of Initiation Factor

38


• Elongation– After initiation complex forms, large

ribosome subunit binds, exposing mRNA codon adjacent to the initiating codon, positioning it for interaction with another amino acid-bearing tRNA molecule.

39


• Translocation– ribosome moves nucleotides along mRNA

molecule

40

A bit about the peptide bond formation

• A peptide bond (or amide bond) is a covalent chemical bond formed between two molecules when the carboxyl group of one molecule reacts with the amine group of the other molecule, thereby releasing a molecule of water.

• This is a dehydration synthesis reaction (also known as a condensation reaction), and usually occurs between amino acids.

• The resulting C(O)NH bond is called a peptide bond, and the resulting molecule is an amide.

• The four-atom functional group -C(=O)NH- is called a peptide link

41

Translation (in more detail)• Termination

– Nonsense codons are recognized by release factors that release the newly made polypeptide from the ribosome.

– There is no tRNA with complimentary antidcodon to (UAA, UAG, UGA)

42

Spliced Gene Transcripts

• DNA sequence specifying a protein is broken into segments (exons) scattered among longer noncoding segments (introns).

• Initially, primary RNA transcript is produced for the entire gene.

– Small nuclear ribonuclearproteins (snRNPs) associate with proteins to form spliceosomes.

Lariat forms, excising introns and splicing exons to form mature mRNA.

alternative splicing

43

Spliceosome

44

RNA Splicing

• During RNA processing, intron sequences are cut out of primary transcript before it is used in polypeptide synthesis.

– remaining sequences are not translated remaining exon sequences are spliced

together to form final processed mRNA

45

Eukaryotic Genes are Fragmented

46

Compartmentalization of processes (thus, transport is important)

DNA Replicationnucleus

Transcription Nucleus

mRNA transferred to cytoplasm

Translation Ribosome (in cytoplasm)

replication

47

From a sequence to a protein

48

An overview of gene expression in eukaryotesCopyright © The McGraw - Hill Companies, Inc. Permission required for reproduction or dis play.

DNA

Nucleus

Primary RNAtranscript

5‘

5‘

3‘

3‘

Nuclearmembrane

In the cell nucleus, RNA polymerase transcribes RNA from DNA.

1

RNA polymerase

49

An overview of gene expression in eukaryotes

50


51


52


53


54

Differences Between Prokaryotic and Eukaryotic Gene Expression

• Most eukaryotic genes possess introns (prokaryotic genes do not.)

• Individual bacterial mRNA molecules often contain transcripts of several genes.

• Eukaryotic mRNA molecules must be completely formed and must pass across the nuclear membrane before translation.

• In prokaryotes, translation begins at the AUG codon preceded by a special nucleotide sequence.

55

Differences Between Prokaryotic and Eukaryotic Gene Expression

• Eukaryotic mRNA molecules have introns cut out and exons joined together before translation.

• Eukaryotic ribosomes are larger than prokaryotic ribosomes.

56

Exceptions to the Central Dogma

DNA

retroviruses use reverse transcriptaseto replicate their genome(David Baltimore and Howard Temin)

RNA viruses

mRNA introns (splicing)(Philip Sharp and Richard Roberts)

RNA editing (deamination of cytosineto yield uracil in mRNA)

RNA interference (RNAi) a mechanismof post-transcriptional gene silencing utilizing double-stranded RNA

RNAs (ribozymes) can catalyze anenzymatic reaction(Thomas Cech and Sidney Altman)

RNA

Protein

Prions are heritable proteins responsiblefor neurological infectious diseases(e.g. scrapie and mad cow) (Stanley Pruisner)

Epigenetic marks, such as patterns ofDNA methylation, can be inherited andprovide information other than the DNAsequence

Nobel Prizes

57

How do mutation effect proteins

• Any change in DNA sequence is called a mutation.• Mutations can be caused by errors in replication,

transcription, cell division, or by external agents

The effects of point mutations• A point mutation is a change in a single base pair

in DNA• A change in a single nitrogenous base can change

the entire structure of a protein because a change in a single amino acid can affect the shape of the protein

58

Normal

Point mutation

mRNA

ProteinStop

Stop

mRNA

Protein

Replace G with A

Mutations – the effects of point mutations

59

Frameshift mutations

• A mutation in which a single base is added or deleted from DNA is called a frameshift mutation because it shifts the reading of codons by one base.

• Structural changes in chromosomes are called chromosomal mutations.

60

• Any agent that can cause a change in DNA is called a mutagen.

• Mutagens include radiation, chemicals, and even high temperatures.

• Forms of radiation, such as X rays, cosmic rays, ultraviolet light, and nuclear radiation, are dangerous mutagens because the energy they contain can damage or break apart DNA.

Causes of Mutations

61

Chromosomal Alterations

• When a part of a chromosome is left out, a deletion occurs.

Deletion

A B C D E F G H

A B C E F G H

62

• When part of a chromatid breaks off and attaches to its sister chromatid, an insertion occurs.

• The result is a duplication of genes on the same chromosome.

Insertion

A B C D E F G H A B C B C D E F G H


63

• When part of a chromosome breaks off and reattaches backwards, an inversion occurs.

Inversion

A B C D E F G H A D C B E F G H


64

• When part of one chromosome breaks off and is added to a different chromosome, a translocation occurs.

A B E FDCBX AWC HGGE HD F

W X Y Z Y ZTranslocation


1 genes expression or genes and how they work: transcription, translation, & more chapter 15

Documents