AP Biology 2007-2008 From Gene to Protein How Genes Work SLIDE SHOW BY KIM FOGLIA (modified) All Blue edged slides are Kim’s (hyperlinks may have been

AP Biology 2007-2008

From Gene to Protein

How Genes Work

SLIDE SHOW BY KIM FOGLIA (modified)All Blue edged slides are Kim’s

(hyperlinks may have been added)

AP Biology

What do genes code for?

proteins cells bodies

How does DNA code for cells & bodies? how are cells and bodies made from the

instructions in DNA

DNA

AP Biology

The “Central Dogma” Flow of genetic information in a cell

How do we move information from DNA to proteins?

_____________

____________

_______________

proteinRNADNA trait

DNA gets all the glory,

but proteins do all the work!

AP Biology

Inheritance of metabolic diseases suggested that genes coded for enzymes each disease (phenotype) is caused by

non-functional gene product lack of an enzyme Tay sachs PKU (phenylketonuria) albinism

Am I just the sum of my proteins?

Metabolism taught us about genes

A B C D E

disease disease disease disease

enzyme 1 enzyme 2 enzyme 3 enzyme 4

metabolic pathway

AP Biology

Beadle & Tatum 1941 | 1958

George Beadle

Edward Tatum

"for their discovery that genes act by regulating definite chemical events"

________________________hypothesis

AP Biology

Wild-typeNeurospora

Minimalmedium

Select one ofthe spores

Grow oncomplete medium

Minimalcontrol

Nucleicacid

CholinePyridoxine Riboflavin Arginine

Minimal media supplemented only with…

ThiamineFolicacid

NiacinInositolp-Aminobenzoic acid

Test on minimalmedium to confirmpresence of mutation

Growth oncompletemedium

X rays or ultraviolet light

asexualspores

spores

Beadle & Tatum

create mutations

positive control

negative control

experimentalsmutatio

n identifi

ed

amino acidsupplements

AP Biology

mRNA

From gene to protein

DNAtranscription

nucleus cytoplasm

aa

aa

aa

aa

aa

aa

aa

aa

aa

aa

aa

proteintranslation

ribosome

trait


Transcription

fromDNA nucleic acid language

toRNA nucleic acid language

AP Biology

RNA ____________ sugar N-bases

_______ instead of thymine U : A C : G

single stranded lots of RNAs

mRNA, tRNA, rRNA, siRNA…

RNADNAtranscription

3 KINDS OF RNA HELP WITH INFO TRANSFER FOR PROTEIN SYNTHESIS

RIBOSOMAL RNA (rRNA)Made in ______________2 subunits (large & small)Combine with proteins to

form ribosomesBacterial ribosomes different

size than eukaryotic ribosomes Evidence for ENDOSYMBIOTIC THEORY Medically significant-some antibiotics target

bacterial ribosomes w/o harming host

rRNA and t-RNA images from Image from: Biology; Miller and Levine; Pearson Education publishing as Prentice Hall; 2006mRNA image from http://wps.prenhall.com/wps/media/tmp/labeling/1140654_dyn.gif


TRANSFER RNA (tRNA)

________________ sequence matches CODON on mRNA to add correctamino acids during protein synthesis

AMINOACYL-tRNA SYNTHETASE Enzyme attaches a specific amino acid using energy from ATP

http://www-math.mit.edu/~lippert/18.417/lectures/01_Intro/


MESSENGER RNA (mRNA)carries code from DNA to ribosomes

AP Biology

Transcription Making mRNA

transcribed DNA strand = _____________ strand untranscribed DNA strand = coding strand

same sequence as RNA synthesis of complementary RNA strand

transcription bubble enzyme

RNA polymerase

template strand

rewinding

mRNA RNA polymerase

unwinding

coding strand

DNAC C

C

C

C

C

C

C

C CC

G

GG

G

G G

G G

G

G

GAA

AA A

A

A

A

A

A A

A

AT

T T

T

T

T

T

T

T T

T

T

U U

5

35

3

3

5build RNA 53

•Transcription

http://bcs.whfreeman.com/thelifewire/content/chp12/1202001.html

AP Biology

RNA polymerases 3 RNA polymerase enzymes

RNA polymerase 1 only transcribes rRNA genes makes ribosomes

RNA polymerase 2 transcribes genes into mRNA

RNA polymerase 3 only transcribes tRNA genes

each has a specific promoter sequence it recognizes

AP Biology

Which gene is read? Promoter region

binding site before beginning of gene TATA box binding site binding site for RNA polymerase

& transcription factors

Enhancer region binding site far

upstream of gene turns transcription

on HIGH

AP Biology

Transcription Factors Initiation complex

transcription factors bind to _____________region suite of proteins which bind to DNA hormones? turn on or off transcription

trigger the binding of RNA polymerase to DNA

•Transcription

AP Biology

Matching bases of DNA & RNA Match RNA bases to DNA

bases on one of the DNA strands

U

A G GGGGGT T A C A C T T T T TC C C CA A

U

UU

U

U

G

G

A

A

A C CRNA

polymerase

C

C

C

C

C

G

G

G

G

A

A

A

AA

5' 3'

AP Biology

Eukaryotic genes have junk! Eukaryotic genes are not continuous

exons = the real gene expressed / coding DNA

introns = the junk inbetween sequence

eukaryotic DNA

exon = coding (expressed) sequence

intron = noncoding (inbetween) sequence

intronscome out!

mRNA’s require EDITING before use

Message in NOT CONTINUOUS INTRONS are removed

Image by Riedell

AP Biology

mRNA splicing

eukaryotic DNA



primary mRNAtranscript

mature mRNAtranscript

pre-mRNA

spliced mRNA

Post-transcriptional processing eukaryotic mRNA needs work after transcription primary transcript = ______________ mRNA splicing

edit out _____________ make mature mRNA transcript

~10,000 bases

~1,000 bases

AP Biology

1977 | 1993

Richard Roberts Philip

SharpCSHL

MITadenovirus

common cold

Discovery of exons/introns

beta-thalassemia

AP Biology

Splicing must be accurate No room for mistakes!

a single base added or lost throws off the reading frame

AUG|CGG|UCC|GAU|AAG|GGC|CAU

AUGCGGCTATGGGUCCGAUAAGGGCCAUAUGCGGUCCGAUAAGGGCCAU

AUG|CGG|GUC|CGA|UAA|GGG|CCA|U

AUGCGGCTATGGGUCCGAUAAGGGCCAUAUGCGGGUCCGAUAAGGGCCAU

Met|Arg|Ser|Asp|Lys|Gly|His

Met|Arg|Val|Arg|STOP|

AP Biology

RNA splicing enzymes

snRNPs

exonexon intron

snRNA

5' 3'

spliceosome

exonexcisedintron

5'

5'

3'

3'

3'

lariat

exonmature mRNA

5'

No, not smurfs!“snurps”

______________ small nuclear RNA proteins

______________ several snRNPs recognize splice

site sequence cut & paste gene

Whoa! I think we just broke

a biological “rule”!

mRNA EDITING

ALL ENZYMES ARE PROTEINS?

______________-RNA molecules that function as enzymes (In some organisms pre-RNA can remove its own introns)

PROCESSING RNA

SPLICEOSOMES

AP Biology

Alternative splicing Alternative mRNAs produced from same gene

when is an intron not an intron… different segments treated as exons

Starting to gethard to

define a gene!

AP Biology

A A AA

A3' poly-A tail

mRNA

5'5' cap

3'

G PPP

50-250 A’s

More post-transcriptional processing Need to protect mRNA on its trip from

nucleus to cytoplasm enzymes in cytoplasm attack mRNA

protect the ends of the molecule add ____________ cap add ____________ tail

longer tail, mRNA lasts longer: produces more protein

AP Biology

mRNA


DNAtranscription

nucleus cytoplasm

aa

aa

aa

aa

aa

aa

aa

aa

aa

aa

aa

ribosome

trait

proteintranslation


Translation

fromnucleic acid language

toamino acid language

AP Biology

How does mRNA code for proteins?

TACGCACATTTACGTACGCGGDNA

AUGCGUGUAAAUGCAUGCGCCmRNA

Met Arg Val Asn Ala Cys Alaprotein

?

How can you code for 20 amino acids with only 4 nucleotide bases (A,U,G,C)?

4

4

20

ATCG

AUCG

AP Biology


mRNA codes for proteins in triplets



Met Arg Val Asn Ala Cys Alaprotein

?

codon

AP Biology

Cracking the code1960 | 1968

Crick determined 3-letter (triplet) codon system

Nirenberg & Khorana

WHYDIDTHEREDBATEATTHEFATRATWHYDIDTHEREDBATEATTHEFATRAT

Nirenberg (47) & Khorana (17) determined mRNA–amino acid match added fabricated mRNA to test tube of

ribosomes, tRNA & amino acids created artificial UUUUU… mRNA found that UUU coded for phenylalanine

AP Biology

1960 | 1968Marshall Nirenberg

Har Khorana

•Determining the code

AP Biology

The code Code for ALL life!

strongest support for a common origin for all life

Code is redundant several codons for

each amino acid 3rd base “wobble”

Start codon AUG methionine

Stop codons UGA, UAA, UAG

Why is thewobble good?

AP Biology

How are the codons matched to amino acids?



aminoacid

tRNA anti-codon

codon

5 3

3 5

3 5

UAC

MetGCA

ArgCAU

Val

AP Biology

mRNA


DNAtranscription

nucleus cytoplasm

aa

aa

aa

aa

aa

aa

aa

aa

aa

aa

aa

ribosome

traitaa

proteintranslation

AP Biology

____________ RNA structure “Clover leaf” structure

anticodon on “clover leaf” end amino acid attached on 3 end

AP Biology

Loading tRNA Aminoacyl tRNA synthetase

enzyme which bonds amino acid to tRNA bond requires energy

ATP AMP bond is unstable so it can release amino acid at ribosome easily

activatingenzyme

anticodontRNATrp binds to UGG

condon of mRNA

Trp Trp Trp

mRNAA C CU GG

C=O

OHOH

H2OO

tRNATrp

tryptophan attached to tRNATrp

C=O

O

C=O

AP Biology

Ribosomes Facilitate coupling of

tRNA anticodon to mRNA codon organelle or enzyme?

Structure ribosomal RNA (rRNA) & proteins 2 subunits

large small E P A

Protein synthesis/quiz

AP Biology

Ribosomes

Met

5'

3'

UUA C

A G

APE

A site (aminoacyl-tRNA site) holds tRNA carrying next amino acid to

be added to chain P site (peptidyl-tRNA site)

holds tRNA carrying growing polypeptide chain

E site (exit site) empty tRNA

leaves ribosome from exit site

Protein synthesis 2

AP Biology

Building a polypeptide ________________

brings together mRNA, ribosome subunits, initiator tRNA

_________________ adding amino acids based on

codon sequence

_________________ end codon 123

Leu

Leu Leu Leu

tRNA

Met MetMet Met

PE AmRNA5' 5' 5' 5'

3' 3' 3'3'

U UA AAACC

CAU UG G

GUU

A AAAC

CC

AU UG GGU

UA

AAAC

CC

AU UG GGU U

A AACCA U UG G

G AC

ValSer

AlaTrp

releasefactor

AA A

CCU UGG 3'

How translation works

AP Biology

Protein targeting ______________________

address label

Destinations: secretion nucleus mitochondria chloroplasts cell membrane cytoplasm etc…start of a secretory pathway

AP Biology

Can you tell the story?

DNA

pre-mRNA

ribosome

tRNA

aminoacids

polypeptide

mature mRNA

5' GTP cap

poly-A tail

large ribosomal subunit

small ribosomal subunit

aminoacyl tRNAsynthetase

E P A

5'

3'

RNA polymerase

exon intron

tRNA

AP Biology AAAAAAAAGTP

20-30b

3'

promoter transcriptionstop

transcriptionstart

introns

The Transcriptional unit (gene?)

transcriptional unit (gene)TAC ACT

DNA

DNATATA5'RNA

polymerase

pre-mRNA

5' 3'

translationstart

translationstop

mature mRNA

5' 3'

UTR UTR

exonsenhancer

1000+b


Protein Synthesis in Prokaryotes

Bacterial chromosome

mRNA

Cell wall

Cellmembrane

Transcription

Psssst…no nucleus!

AP Biology

Prokaryote vs. Eukaryote genes Prokaryotes

DNA in cytoplasm circular

chromosome naked DNA

no introns

Eukaryotes DNA in nucleus linear

chromosomes DNA wound on

histone proteins introns vs. exons

eukaryoticDNA



intronscome out!

AP Biology

Transcription & translation are simultaneous in bacteria DNA is in

cytoplasm no mRNA

editing ribosomes

read mRNA as it is being transcribed

Translation in Prokaryotes

AP Biology

Translation: prokaryotes vs. eukaryotes Differences between prokaryotes &

eukaryotes time & physical separation between

processes takes eukaryote ~1 hour

from DNA to protein no RNA processing

SEE PROCESSING VIDEO

COMPLETING PROTEINS POLYRIBOSOMES (POLYSOMES)

Numerous ribosomes translate same mRNA at same time

3-D folding (1’, 2’, 3’ structure) Chaparonins

POST-TRANSLATIONAL MODIFICATIONS Some amino acids modified by addition of

sugars, lipids, phosphate groups, etc Enzymes can modify ends, cleave into pieces

join polypeptide strands (4’ structure)

Ex: Made as proinsulin then cut

Final insulin hormonemade of two chains connected by disulfide bridges

http://www.vivo.colostate.edu/hbooks/pathphys/endocrine/pancreas/insulin.html

1st to suggest genes dictate phenotypes through enzymes that catalyze specific chemical reactions

Postulated that the symptoms of an inherited disease are due to inability to make a specific enzyme

Coined term “inborn errors of metabolism” to describe such diseases Beginning of “One gene-one enzyme” hypothesis ALCAPTONURIA- “black urine” disease- defect in enzyme that breaks down amino acid

tyrosine

http://www.personal.psu.edu/faculty/w/x/wxm15/Online/Molecular%20Biology/media/phenylalanine.gifhttp://www.nature.com/bjp/journal/v147/n1s/images/0706466f5.jpg

ARCHIBALD GARROD1902

Mutations Point mutations

single base change base-pair

substitution silent mutation

no amino acid change redundancy in code

missense change amino acid

nonsense change to stop codon

Slide from Explore Biology by Kim Foglia

Point mutation leads to Sickle cell anemia

What kind of mutation?


Sickle cell anemia


Mutations

Frameshift shift in the reading

frame changes everything

“downstream” insertions

adding base(s) deletions

losing base(s) More damaging at

beginning of gene than at end

Slide modified from: Explore Biology by Kim Foglia

DNA → DNA ____________

DNA → RNA ____________

RNA→ Protein ___________

WHAT IS A “GENE”?

Mendel’s factors determine phenotype

T.H. Morgan- genes located on specific chromosomes

Beadle and Tatum’s “one gene-one enzyme”

Became “One gene-one polypeptide”

- Some proteins made of more than one polypeptide chainEx: hemoglobin has 4 polypeptide chains

Now: “one gene – one polypeptide or RNA” - Not all genes code for proteins

AP Biology

Can you tell the story?


Any Questions??

What color would a smurf turnif he held his breath?

Documents

AP Biology 2007-2008 From Gene to Protein How Genes Work SLIDE SHOW BY KIM FOGLIA (modified) All Blue edged slides are Kim’s (hyperlinks may have been