TRANSLATION

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TRANSLATIONINTRODUCTION

Protein (amino acid polymer, N terminus, C terminusRibosomes: 30 S subunit plus 50 S subunit form functional 70 S ribosomemRNA is read 5' to 3'Initiation Codons; Stop CodonsPolysomes

INITIATIONInitiator tRNA (N - formyl methionyl tRNA) Initiation complex

ELONGATIONPeptidyl transferaseA, P, E sites on ribosome

TERMINATIONStop codons

SHINE-DALGARNO

2

3AMIDES

ACID

AMINE

AMIDE

PEPTIDE BOND

PEPTIDE-AMIDE FORMED BETWEEN TWO AMINO ACIDS: ALANYL-GLYCINE

MONO SUBSTITUTED DISUBSTITUTED

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5'

3'

3'

5'

TRANSCRIPTION

TRANSLATION

UNTRANSLATED REGION

5' 3'

N CCODENOBELHOLLEY

NIRENBERGKHORANA

[STOP CODONS = UAA/UAG/UGA]

GENEPROMOTER

+1

5RIBOSOMES - NON-SPECIFICTRANSLATE ANY MESSAGEPROKARYOTIC

70S 50S + 30SSUBUNITS

COMPOSITION 50S 30S

NUMBER OF DIFFERENT PROTEINS 34 21

NUMBER OF DIFFERENT rRNA MOLECULES

ONE - 5SONE - 23S ONE 16S

POLYSOMES

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CODON FOR PHENYLALANINE

tRNA - IS AN ADAPTER - ADAPTS AMINO ACID AND CODON

ANTICODON3' 5'

mRNA

5' 3'

TRANSFER RNA tRNA ~ 60EACH SPECIFIC FOR 1 AMINO ACID AND 1 CODON

HO5'

3'

tRNAPHE

SPECIFIC FORPHENYLALANINE

8OVERALL:

AMINO ACID + ATP + tRNA AMINO ACYL-tRNA + AMP + PPiSTEP 1:

NUCLEOTIDYLTRANSFER

PPi

AMINO ACID + ATP AMINO ACYL-AMP + PPiAMINO ACID ACTIVATING ENZYME =

AMINO ACYL~tRNA SYNTHETASE

+

ANHYDRIDE

9STEP 2: AMINO ACYL AMP + tRNA AMINO ACYL~tRNA + AMP

3' ENDAMINO ACYL~AMP

tRNA

AMINOACYL~tRNA AMP

7GENETIC CODE IS DEGENERATE -

DIFFERENT TRIPLETS ENCODE THE SAME AMINO ACID

"WOBBLE" MEANS THAT SOME SPECIFIC tRNAs CAN READ MORE THAN ONE CODON

"WOBBLE" MAKES IT POSSIBLE FOR AN ORGANISM TO TRANSLATE ALL 61 CODONS WITH FEWER THAN 61 tRNAsPROKARYOTES OFTEN HAVE 60 - 80 tRNAs

SOME DIFFERENT tRNAs CARRY THE SAME AMINO ACIDSSOME HIGHLY SPECIALIZED PROKARYOTES HAVE ~ 40

tRNA GENES

ACAACGACCACU

ALL CODE THREONINE

A THREONINE tRNA WITH AN ANTI-CODON OF3' UGG 5'

5' ACC 3'5' ACU 3'

PAIRS WITHOR THREONINE

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FORMYL METHIONINE~tRNA[SPECIFIC FOR STARTING TRANSLATION]

INITIATING TRANSLATION

METHIONINE + tRNAfMET

+ ATP+ ACTIVATING ENZYME

fMET

METHIONYLtRNAfMET

fMET

FORMYLGROUP

[FOLIC ACID]

N-FORMYL METHIONYL

~tRNAfMET

10INITIATION COMPLEXES

F MET

F MET

pppG5' 3'

mRNA + 30S + F - MET~tRNAfMET

+ GTP + INITIATION PROTEINS

30S SUBUNIT30S INITIATION

COMPLEX

50S SUBUNIT

E P A

mRNA

EXITPEPTIDYL

ACCEPTORpppG 3'

70S RIBOSOME

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PEPTIDYL TRANSFERASE

F-METSER

F-MET SER TYR

P A

ELONGATION-POLYMERIZATION

[CHLORAMPHENICOL]

5' G

5' G

3'

3'P A

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TRANSLOCATION [ERYTHROMYCIN – INHIBITS MOVEMENTAMINO ACYL TRNA FROM A SITE TO P SITE, BLOCKS ELONGATION;AZITHROMYCIN]

F-METSER TYR

P A5' 3'

P A

TYRF-MET

SER

5' 3'

HO

HO

E

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P

TYRF-MET SER

5'

HO

AE

GLNHO

GUC

ALL INHIBIT 30S RIBOSOME FUNCTION

TETRACYCLINE – INHIBITS AMINO ACYL TRNA BINDING A SITEDOXYCYCLINE - NEWER

STREPTOMYCIN - TRNA MISMATCHING, MIS-TRANSLATION SPECTINOMYCIN - BINDS 30S SUBUNIT; EXACT MECHANISM ?

PEPTIDYL TRANSFERASE

PEPTIDYLTRANSFERASE

UN-CHARGEDtRNA PEPTIDYL ~ tRNA

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15HOW DO RIBOSOMES KNOW WHERE TO START

TRANSLATING?

A. HOW TO DISTINGUISH INITIAL AUG FROM INTERNAL AUG

B. HOW TO RECOGNIZE SEVERAL INITIATING AUG's IN ONE mRNA

AUG

AUG UAA5' 3'

PROTEIN CODING REGIONINTERNAL AUG

UNTRANSLATEDREGION

5' 3'AUG

AUG

1st CODINGREGION

2nd CODINGREGION

3rd CODINGREGION

AUG

POLY CISTRONIC mRNA RIBOSOMES TRANSLATE ALL THREE INDEPENDENTLY.

HOW?

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PROTEIN CODING REGIONS ARE PRECEDED BY SHINE-DALGARNO SEQUENCES [PROKARYOTES]

5' 3'

5'

CODING REGION

AUGCODING REGION

~ 7+1

SHINEDALGARNO

SHINE DALGARNO SEQUENCES:

SHINE

DALGARNOSHIN

E

DALGARNO

AUG

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30S SUBUNIT

SITES

BINDING BETWEEN SHINE-DALGARNO AND rRNA (16S) POSITIONS mRNA SO THAT AUG

OCCUPIES P SITE

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mRNA

PROTEINSRIBOSOMES

REPLICATION, TRANSCRIPTION & TRANSLATION - DYNAMIC

THEY ALLGO ON ATTHE SAME

TIME !

19RIBOSOMES ARE RIBOZYMES - THE ACTIVE SITE FOR CATALYZING PEPTIDE BOND

FORMATION IS ENTIRELY RNA

23S rRNA IS PEPTIDYL TRANSFERASEADENINE [#2451] PARTICIPATES IN FORMING PEPTIDE

BONDSADENINE #2451

H BOND

ADENINE N3 TAKES PROTONNITROGEN ATTACKS

APPRECIATE !

CHLORAMPHENICOLBINDS A 2451 &A 2452 - INHIBITS

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H BOND STABILIZES

INTERMEDIATE

tRNA OXYGEN TAKES PROTONtRNA PEPTIDE BOND BREAKS

OXYANION ELECTRONS FORM C O

APPRECIATE !

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PEPTIDE BOND FORMS

GROWING PEPTIDE HAS BEEN ADDED TO ONE MOREAMINO ACID

OR; ONE MORE AMINO ACID HAS BEEN ADDED TO GROWING PEPTIDE

ADENINE #2451 IS NOW READY TO REPEAT THE PROCESS

APPRECIATE !