Translation : Introduction

Molecular Biology

Dr. Aga Syed Sameer

CSIR Lecturer (Demonstrator)Department of Biochemistry,Medical College,Sher-I-Kashmir Institute of Medical Sciences, Bemina, Srinagar, Kashmir, 190018. India.

Translation is the conversion of the information in the nucleic acid sequence to polypeptides of a specific amino acid sequence

The process of translation requires a system to bring the mRNA together with the translating molecules (tRNAs)

The system must also catalyze the polymerization of the amino acids into a polypeptide sequence

The message in mRNAs is the sequence of nucleotide bases and is always read in the 5' to 3' direction

The polypeptide chain is made starting with the amino end of the chain first and carboxyl end of the chain last

Thus, the 5' end of the coding region of the mRNA contains the code for the amino end of the polypeptide chain

TranslationProcess

Genetic code actually consists of linear sequence of the nucleotides that make up the DNA or RNA

Within the linear sequence there is a set of triplet nucleotides each of which called as codon specifies an amino acid

As there are four different nucleotides that make up the DNA or RNA, thus, there are 43=64 codons that constitute a genetic code

Genetic CodeFeatures

Out of 64 codons only 61 called as sense codons specify the amino acids during the translation process

Out of 61 codons, AUG called as initiator codon signals the beginning of the polypeptide and codes for methionine (or N-formyl-methionine)

Three codons called as termination codons- UAA (ochre), UAG (Amber) & UGA (Opal) normally signal the end of polypeptide synthesis

Five amino acids each of which is coded by four different codons and three for each of which there are six codons. This phenomenon in which an amino acid is coded by more than one codon is called as degeneracy

The third base of the codon, which base pairs with the first base of anti-codon of tRNA (5'→3'), does not confer the coding specifity in most cases. This phenomenon is called as ambiguity. As a result of which only 32-35 tRNAsare required to translate all 61 codons

Genetic CodeFeatures

This hypothesis was first of all given by Crick to explain the ability of the tRNA molecule to recognise the multiple codons and thus to acts as the adaptor for the many codons

Wobble in literal terms means that the interaction between the codon of the mRNA and the anticodon of the tRNA is not strictly specific

It occurs because of the non-standard hydrogen interactions of two kinds:

GU Interactions:

Inosine Interaction with A, U & C:

Wobble HypothesisWhat is it all about?

During an interaction of the codon and anticodon, guanine is able to base pair with the uracil.

Thus if tRNA anticodon is 3׳-XXG-5׳ it can base pair with 5׳-XXU-3׳ and 5׳-XXC-3׳

And, if the anticodon is 3׳-XXU-׳5 it can base pair with 5׳-XXA-3׳and 5׳-XXG-3׳

Inosine when present in the anticodon at the third position i.e., 3׳-XXI-5 ׳can base pair with the 5׳-XXA-3, 5׳-XXU-3׳ -׳5 &XXC-3׳

Wobble HypothesisInteractions

In humans GU interaction is used in eight tRNA’s but in every case anticodon is 3׳-XXG-5׳;

Anticodon with sequence 3׳-XXU-5׳ is not used in eukaryotes

Also eight human tRNA’s have anticodons having sequence ׳XXI-5-׳3 but these decode only 5׳-XXU-3׳ ׳XXC-3-׳5 &codons

Wobble HypothesisInteractions

The process of translation takes place on the ribosomes which act as the seats for the decoding of the information in mRNA into the sequence of amino acids

In prokaryotes, ribosomes have three sites each having a characteristic function associated with it and each contributed by both the 30S & 50S subunits

A or aminoacyl site which accepts the aminoacyl-tRNAsfrom the elongation phase of the translation

P or peptidyl site which has the function to catalyse the formation of the peptide bond between the proceeding and succeeding amino acids by the peptidyltransferase activity that is inherent to it, and

E or exit site( present in large subunit only)

In eukaryotes only first two sites are present

Translation Aspects

In prokaryotes translation start site is defined by the conserved sequence located 8-13 bases upstream of the first codon to be translated

Called as Shine-Dalgarno sequence is a purine rich sequence of 5׳-AGGAGGU-3׳

These sequence base pairs with a complementary sequence of 5׳-ACCUCCU-3׳ located on the 3׳ end of the 16S rRNA in the small subunit of the ribosome

During translation of the mRNA molecule it is not only one ribosome that translates the sequence but there occurs a cluster of 10 to 100 ribosomes that translate the mRNA sequence into protein simultaneously by binding mRNA one after the other. This assembly of the ribosomes on the single mRNA molecule is called as polysomes

Translation Aspects

As there are only 20 amino acids which are normally found in the proteins coded by 61 codons

But, there are about 30-45 tRNAs in prokaryotes and some 50 in eukaryotes

This means that there are many tRNAs which carry same amino acid bound to their amino acid (3’) accepting arm

These are called as isoaccepting tRNAs

Binding or transfer of amino acid to the adenosine moiety of the 5’-CCA-3’ amino acid accepting arm of tRNA is called as the charging of the tRNA

It is bought about by the highly conserved class of enzymes called as aminoacyl-tRNA synthetases, of which there are two types: I & II

Translation Charging of tRNA

Each enzyme is specific for one amino acid and one or more corresponding tRNAs

Type I enzyme catalyses the transfer of amino acid to the 2’-OH group of the adenosine moiety located at the CCA-3’ end

Type II enzyme catalyses the transfer of amino acid to the 3-OH group of the adenosine

Both of these enzymes catalyse the charging of tRNA in two steps

In the first step carboxyl group of the amino acid reacts with the α–phosphoryl group of ATP to form an anhydride linkage so as to produce enzyme-bound intermediate aminoacyl adenylate

In the second step aminoacyl group is transferred to its corresponding specific tRNA.

Thus overall process involves the consumption of energy equivalent to two ATPs

Translation Charging of tRNA

mRNA is read in the 53→׳ direction ׳

Ribosomes began translating the 5׳- end of the mRNA before the transcription is complete (in prokaryotes)

Thus in other words protein synthesis begins at the amino-terminal

The AUG initiator codon specifies the amino-terminal methionine residue

In both pro & eukaryotes there are two different tRNAs that code for the AUG codon

Translation The First Codon

One is used when AUG is a initiator codon where it incorporates N-formyl methionine (in prokaryotes) and methionine (in eukaryotes)

Other is used to incorporate methionine in response to the AUG located in the internal position in mRNA

They are respectively designated as tRNAfMet & tRNAMet

Both the tRNAs are charged by same enzyme Met-tRNA synthetase

But methionine bound to tRNAfMet is then modified by addition of formyl group by transformylase to give N-formylmethionyl-tRNAfMet

Translation The First Codon

A reading frame without a termination codon among 50 or more codons is referred to as an open reading frame (ORF)

In a random sequence of nucleotides, 1 in every 20 codons in each reading frame is, on average, a termination codon

Translation ORFs

Interaction

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