Chapter 14

Gene Activity: How Genes Work

14.1 The Function of GenesGenes are segments of DNA that specify amino acids in a protein

Sir Archibald Garrod proposed a link between genes and proteins

Genes Specify EnzymesBeadle and Tatum (1940) experimented with Neurospora (mold similar to Sordaria)

they induced mutations that made the mold only able to grow on enriched medium

Fig. 14.1 Beadle & Tatum experiment

One gene, one enzyme experiment

14.1 The Function of Genes Genes Specify Enzymes, cont.

Beadle and Tatum (1940), cont.concluded that mold must have been missing an enzyme needed to produce missing nutrient

therefore, they proposed the “one gene, one enzyme” hypothesis

14.1 The Function of Genes Genes Specify Enzymes, cont.

Pauling and Itano (1949)demonstrated that sickle cell hemoglobin is different from normal hemoglobin using electrophoresis

showed that a mutation in DNA resulted in a change in the structure of a protein

“one gene, one enzyme” revised to “one gene, one polypeptide”

Fig. 14.2 Sickle cell disease

Fig. 14.2c R group and properties

14.1 The Function of Genes From DNA to RNA to Protein

gene: segment of DNA that specifies the amino acid sequence of a protein

genes pass information to RNA (ribonucleic acid), which is more directly responsible for building proteins

14.1 The Function of Genes DNA to RNA to Protein, cont.

RNA differs from DNA

Fig. 14.3 RNA structure

14.1 The Function of Genes DNA to RNA to Protein, cont.

there are three major classes of RNA, each with its own size, shape, and function

messenger RNA (mRNA): takes message from DNA to ribosomes

transfer RNA (tRNA): carries amino acids to ribosomes

ribosomal RNA (rRNA): makes up ribosomes (along with ribosomal proteins)

14.1 The Function of Genes DNA to RNA to Protein, cont.

steps of protein synthesis (gene expression)

transcription: DNA serves as template to make RNA

translation: mRNA transcript codes for sequence of amino acids

this is Crick’s Central Dogma of Molecular Biology

Overview of gene expression

Overview of gene expression

14.2 The Genetic Code

The genetic code is a triplet codeeach codon consists of three nucleotide bases and codes for one amino acids

three nucleotides is necessary to code for 20 amino acids

14.2 The Genetic Code Finding the Genetic Code

Nirenberg and Matthei (1961) used an enzyme to produce an RNA molecule made entirely of uracil

a protein composed only of phenylalanine resulted from the RNA, therefore UUU = phe

in a similar way, the other amino acids were assigned mRNA codons

14.2 The Genetic Code Finding the Genetic Code

Nirenberg and Matthei (1961)all uracil RNA made a protein that was all phenylalanine

therefore UUU = phein a similar way, the other amino acids were assigned mRNA codons

Fig. 14.5 Messenger RNA codons

14.2 The Genetic Code Finding the Genetic Code, cont.

properties of the genetic code1. it is degenerate (redundant); most amino acids have more than one codon

2. it is unambiguous; each codon has only one meaning

3. it has start and stop signals start codon is AUG; codes for MET UAA, UAG, UGA stop translation

(no amino acid encoded)

The genetic code (easier to read)

14.2 The Genetic Code The Code Is Universal

the code is used by all living thingsevidence for the common ancestry of living things

there are some exceptions to the universality, but they generally involve just a few codons

Overview of transcription

14.3 First Step: Transcription

During transcription, a segment of DNA serves as a template for the production of RNA Messenger RNA Is Formed

initiationa promoter signals the start of a gene, direction of transcription, strand to be transcribed

promoters are specific nucleotide sequences on DNA (often TATA...)

attract RNA polymerase

Initiation of transcription

14.3 First Step: Transcription Messenger RNA Is Formed, cont.

initiation, cont.transcription factors bind to the promoter and help RNA polymerase recognize and bind to DNA

elongationRNA polymerase opens DNA strands and synthesizes mRNA along the template strand

14.3 First Step: Transcription Messenger RNA Is Formed, cont.

elongation, cont.RNA polymerase, cont.

reads 3’-5’ makes mRNA 5’-3’ mRNA grows at a rate of 30-60

nucleotides/secondonly newest portion of mRNA is bound to DNA; most of the new strand dangles off to the side

Fig. 14.6 Transcription

14.3 First Step: Transcription Messenger RNA Is Formed, cont.

elongation, cont.many RNA polymerase molecules might be working at the same time

template/non-template strands of DNA attach back together after RNA polymerase passes

Numerous RNA transcripts

Fig. 14.7b RNA polymerase

14.3 First Step: Transcription Messenger RNA Is Formed, cont.

terminationspecific nucleotide sequences signal “stop” and cause RNA polymerase to release DNA

“stop” sequence is called a terminator, usually AAUAAA on the mRNA in eukaryotes

(do not confuse this with a stop codon)

the product is primary RNA

Transcription animation

14.3 First Step: Transcription RNA Molecules Are Processed

occurs before leaving nucleus5’ cap added to tell ribosome where to attach

3’ poly-A tail added to aid transport out of nucleus and inhibit degradation

introns removed by spliceosomesintrons: in between exons; 95%exons: coding regions that will be expressed

RNA processing

RNA splicing

End result of splicing

Prokaryotes versus eukaryotes

14.4 Second Step: Translation

During translation, the mRNA message is translated to an amino acid sequence The Role of Transfer RNA

tRNA molecules transfer amino acids to ribosomes

tRNA has an anticodon that is complementary to a codon

tRNA also has an amino acid binding site, making it “bilingual”

Fig. 14.9 Structure of transfer RNA

14.4 Second Step: Translation The Role of Transfer RNA, cont.

most cells have 40 different tRNA molecules

this is less than the 61 amino-acid-coding codons and more than the 20 different amino acids

perhaps some tRNAs can pair with more than one codon as long as first two positions are correct

called wobble hypothesis

14.4 Second Step: Translation The Role of Transfer RNA, cont.

aminoacyl-tRNA synthetases attach correct amino acid to each tRNA

requires ATP20 different types (1 for each amino acid)

this means they can accommodate different kinds of tRNA

14.4 Second Step: Translation The Role of Ribosomal RNA

structure of a ribosomeconsists of a small and large subunit

made in nucleolus60% rRNA, 40% proteinhas 3 binding sites for tRNA

A site: amino acid site P site: peptide site E site: exit site

has 1 binding site for mRNA

Fig. 14.10a,b Ribosome structure

14.4 Second Step: Translation The Role of Ribosomal RNA, cont.

function of a ribosometRNA binding sites facilitate base pairing between tRNA anticodons and mRNA codons

rRNA joins amino acids as a polypeptide is synthesized

multiple ribosomes can translate same mRNA simultaneously; complex called a polyribosome

Fig. 14.10c,d Polyribosome

14.4 Second Step: Translation Translation Requires Three Steps

initiation (in prokaryotes)brings all the translation components together

small ribosomal subunit attaches to mRNA at 5' end at initiation sequence (AUG)

tRNA with anticodon UAC (carrying amino acid MET) attaches to initiator

Fig. 14.11 Initiation

14.4 Second Step: Translation Translation Requires Three Steps

initiation, cont.tRNA, mRNA, MET and small subunit form initiation complex

large ribosomal subunit attaches to initiation complex

tRNA is at P siteA site is for next tRNAE site is for tRNAs leaving ribosome

Fig. 14.12 Elongation

14.4 Second Step: Translation Translation Requires Three Steps

elongationpolypeptide increases in length one amino acid at a time

next tRNA with complementary anticodon binds to A site

peptide transferred to this tRNA as ribozyme joins the amino acids at A and P site (forms peptide bond)

14.4 Second Step: Translation Translation Requires Three Steps

elongation, cont.ribosome moves forward

tRNA at A site (with peptide) shifted to P site

tRNA at P site (“empty”) shifted to E site and exits

the next tRNA binds at A site and process repeats

14.4 Second Step: Translation Translation Requires Three Steps

terminationtranslation components separateoccurs at a stop codon (UAG, UAA, UGA)

no tRNA with anticodon for theserelease factors cleave polypeptide from last tRNA and cause ribosome to split, releasing mRNA and new protein

protein is properly folded

Fig. 14.13 Termination

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation

Translation animation

Fig. 14.14 Summary

Nucleus

DNA

Replication

RNA

Transcription

Central Dogma of Molecular Biology

Protein

Translation Enzymes

StructuralComponentsof the Cell

Cytoplasm (ribosomes)