molecular biology lecture 6

8/12/2019 Molecular Biology Lecture 6

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BIOL321 - Translation

Cristofre MartinDepartment of BiochemistrySt. George’s University



Central Dogma of Genetics and Molecular

Biology

DNA RNA ProteinTranscription Translation

Reverse transcription

Replication



General structural formula for an amino acid

The structure of allamino acids consistsof a central carbon atom to which is

bonded an aminogroup , a carboxylgroup , and ahydrogen atom (exception proline).



Bound to thecentral carbon is

the variable Rgroup whichdefines eachdifferent aminoacid species .

The R groupsdefines thespecific

properties of the

polypeptide.



Amino acids of a polypeptide chain are joined by apeptide bond formed between the carboxyl group of oneamino acid and the amino group of an adjacent amino acid.

The N-terminal end defines the beginning of the polypeptide chain and the C-terminal defines the end of

the polypeptide chain.



Peter J. Russell, iGenetics : Copyright © Pearson Education, Inc., publishing as Benjamin Cummings.

There are four different types of amino acids:



Marshall Nirenburg

In 1968, Marshall Nirenburg received the Nobel Prize fordeciphering the geneticcode.

Shared the award withRobert W. Holley and HarGobind Khorana



Nirenburg used synthetic mRNAs and in vitro translation systemsto decipher the genetic code.

For example,

a synthetic mRNA with the sequence UUUUUUUUUUUU

translated to the polypeptide: Phe Phe Phe Phe

or

a synthetic mRNA with the sequence UCUCUCUCUCUC

translated to the polypeptide: serine-leucine-serine-leucine


http://slidepdf.com/reader/full/molecular-biology-lecture-6 9/33Peter J. Russell, iGenetics : Copyright © Pearson Education, Inc., publishing as Benjamin Cummings.

The Genetic Code

Characteristics of theGenetic Code:

1. The code is a tripletcode. Each mRNAcodon that specifies anamino acid in a

polypeptide chainconsists of threenucleotides.



Characteristics of the Genetic Code (cont.)

2. The code is comma free . The mRNA is readcontinuously, three nucleotides at a time, withoutskipping any nucleotides of the message.

Normal mRNA: AUG ACA CAU AAC GGC UUC GUA

Amino acids: Met Thr His Asn Gly Phe Val

translation

3. The code is nonoverlapping . The mRNA isread in successive groups of three nucleotides.



Characteristics of the Genetic Code (cont.):

4. The code is almost universal . Almost allorganisms found on the earth use the same

genetic language.

**Exceptions - mitochondria of some

organisms and the genomes of the protozoanTetrahymena have minor changes to the code.



Characteristics of the

Genetic Code (cont.):

5. The code is degenerate .

With two exceptions, (onlyAUG codes for methionineand only UGG codes fortryptophan), more one codonoccurs for each amino acid.



Characteristics of the Genetic Code (cont.):

6. The code has start and stop codon signals .

AUG (which codes for methionine) is almost

always the start codon for protein synthesis.

Three other codons do not specify an amino

acid, but are stop codons indicating thetermination of the translation process. Thesecodons are UAG (amber), UAA (ochre), andUGA (opal).



Base-pairing wobble Problem :

Sometimes cells donot carry tRNAs forall codons.

Solution : Wobble isa property thatallows an identicaltRNA to bind todifferent codons atthe third base.



The properties of the genetic code (atriplet reading frame) make it such that asmall change in the protein coding

sequence can have profound effects onthe resulting polypeptide sequence.




Reversion of a deletion frameshift mutation by anearby addition mutation.




Hypothetical example showing how three nearby(addition) mutations restore the reading frame, givingnormal or near normal function.



• Silent mutation: » Changed codon may code for same amino acid» UCA UCU same amino acid, Serine» Called “silent” mutation

• Missense mutation: » Changed codon may code for different amino acid» UCA CCA = Ser Pro

• Substitution of different amino acid missense mutation

• Nonsense mutation: » UCA UAA = Ser termination codon» Causes premature termination of sequence =

nonsense mutation

Consequences of altering the nucleotide sequence: Changing a single base in mRNA sequence = point mutation 3

possible results



Consequences of

altering thenucleotidesequence:



Molecular details of the attachment of an aminoacid to a tRNA molecule.

1) The anticodon is thesequence of threenucleotides that base-pairswith a codon in mRNA.

2) The amino acid matching

the codon/anticodon pair isattached to the 3’ end of thetRNA.



The various structure oftRNA molecules.



Base-pairing wobble Problem :

Sometimes cells donot carry tRNAs forall codons.

Solution : Wobble isa property thatallows an identicaltRNA to bind todifferent codons atthe third base.



Charging of a tRNA molecule by aminoacyl-tRNA synthetase toproduce an aminoacyl-tRNA (charged tRNA).

1) There is a different

aminoacyl-tRNA synthetasefor each amino acid.

2) The amino acid is activatedthrough linkage of AMP to itscarboxyl group formingaminoacyl-AMP (via ATPhydrolysis).

3) The enzyme then transfers thecarboxyl group of the aminoacid from AMP to thehydroxyl group at the 3’ endof the tRNA forming the finalaminoacyl-tRNA.



The Ribosome is the Translation Machine

A ribosome contains three bindingsites for RNA molecules: one is forthe mRNA and two (called the A-site,and the P-site) are for tRNAs.

The A- and P- sites are close enough

together for their two tRNAmolecules to be forced to form base

pairs with adjacent codons on themRNA - thus maintaining the correct

reading frame on the mRNA.



The Ribosome

1) The ribosome has twosubunits each composed ofrRNA and numerous

proteins.

2) In E.coli the small 30S

subunit is involved in theinitial binding to the mRNA.

3) The larger 50S subunit

contributes the peptidyltransferase activity.

4) Both the 30S and 50Scontribute to the P- and A-sites.



Initiation of protein synthesis in prokaryotes

A 30S ribosomal

subunit, complexedwith initiation factors and GTP binds tomRNA and fMetinitiator tRNA to forma 30S initiationcomplex .

The 30S ribosome

binds to a specificmRNA sequencecalled the Shine-Dalgarno sequence(typically5’ - AGGAGG - 3’).

The fMet initiator tRNA binds to thestart codon AUG.



The 50S ribosomalsubunit binds, forming a70S initiation complexduring which time theinitiation factors arereleased and GTP ishydrolyzed.

In eukaryotes , the smallribosomal subunit first

binds to the methylatedcap at the 5’ end of themRNA. It then migratesto the initiation site -usually the first AUG itencounters.



1) A group of proteins called elongation factors (EF-Tu) and GTPusher the appropriate tRNA into the A site of the ribosome.

2) Peptide bond formation between the two adjacent amino acids by

peptidyl transferase.3) The formation of the first peptide bond causes the initiating tRNA

in the P site to be release its amino acid and dissociates from themRNA.

4) The ribosome moves along to the next codon (requires elongationfactors and GTP).



Peter J. Russell, iGenetics : Copyright © Pearson Education, Inc., publishing as Benjamin Cummings.Peter J. Russell, iGenetics : Copyright © Pearson Education, Inc., publishing as Benjamin Cummings.

The formation of a peptide bond between the first two aminoacids (fMet and Ser) of a polypeptide chain is catalyzed onthe ribosome by peptidyl transferase.




A polysome - a number of ribosomes each translating thesame mRNA sequentially.



Micrograph of a group of polyribosomes.

The green arrow indicated theDNA strand, and the red arror

indicated a polyribosome that istranslating an mRNA moleculeinto protein.



Termination of translation

Translation termination is facilitated by the association of releasing factors with thestop codons (UAG, UAA and UGA).

Eukaryotes possess a single releasing factor - eRF.

Prokaryotes possess three releasing factors: RF1 (recognizes UAA and UAG), RF2(recognizes UAA and UGA) and RF3 which stimulates termination.

Peptidyltransferase releases polypeptide from tRNA in the P site.

molecular biology lecture 6

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