The genetic code and translation Dr.Aida Fadhel Biawi 2013

DNA RNA POLYPEPTIDE

Virtually all organisms share the same genetic code! Just like we have 26 letters in the alphabet to construct words, the alphabet of DNA has 4 letters to use to construct polypeptides. So how is the code used to construct polypeptides? DNA template C A G T A A G C C RNA strand G U C A U U C G G

In a metabolically active cell, approximately 3% to 5% of the cellular RNA is mRNA, 90% is rRNA, and about 4% is tRNA. -Hundreds of different mRNAs can be in one cell. -By contrast, there are four types of rRNA. Three of the rRNAs combine with a set of proteins to form a ribonucleoprotein complex called the large ribosomal subunit. The other rRNA combines with another set of proteins to form a small ribosomal subunit. In the cytoplasm of the cell, one large and one small ribosomal subunit combine to form a ribosome. The ribosome is the site of protein synthesis, and an active cell can have thousands of ribosomes.

There are approximately 50 different types of tRNA molecules in a cell that is actively synthesizing protein. The tRNAs range in length from about 75 to 93 nucleotides. An amino acid is linked enzymatically by its carboxyl end to the 3' end of a specific tRNA.

Three major types of RNA are transcribed. mRNA (messenger RNA) - encodes genetic information from DNA & carries it into the cytoplasm. Each three consecutive mRNA bases forms a genetic code word (codon) that codes for a particular amino acid. 5’3’ Start codon

rRNA (ribosomal RNA) - associates with proteins to form ribosomes. Subunits are separate in the cytoplasm, but join during protein synthesis (translation). large subunit small subunit

Functionally competent ribosomes Ribosomes are large complexes of protein and ribosomal RNA. They consist of two subunits—one large and one small—whose relative sizes are generally given in terms of their sedimentation coefficients, or S (Svedberg) values. The prokaryotic 50S and 30S ribosomal subunits together form a 70S ribosome. The eukaryotic 60S and 40S subunits form an 80S ribosome. Prokaryotic and eukaryotic ribosomes are similar in structure, and serve the same function, namely, as the “factories” in which the synthesis of proteins occurs. -The large ribosomal subunit catalyzes formation of the peptide bonds that link amino acid residues in a protein. -The small subunit binds mRNA and is responsible for the accuracy of translation by ensuring correct base-pairing between the codon in the mRNA and the anticodon of the tRNA.

tRNA (transfer RNA) - transports specific amino acids to ribosome during protein synthesis (translation). Anticodon - specific sequence of 3 nucleotides; complementary to an mRNA codon. Amino acid accepting end Anticodon sequence determines the specific amino acid that binds to tRNA.

Hydrogen bond Amino acid attachment site RNA polynucleotide chain Anticodon

tRNA

This decoding process requires two types of adapter molecules: tRNAs and enzymes called aminoacyl- tRNA synthetases. First we describe the role of tRNAs in decoding mRNA codons, and then examine how synthetases recognize tRNAs.

All tRNAs have two functions: 1- to be chemically linked to a particular amino acidamino acid basebase-pair 2- and to base-pair with a codon in mRNA so that the amino acid can be added to a growing peptide chain.basecodonpeptide Each tRNA molecule is recognized by one and only one of the 20 aminoacyl-tRNA synthetases. Likewise, each of these enzymes links one and only one of the 20 amino acids to a particular tRNA, forming an aminoacyl- tRNA. Once its correct amino acid is attached, a tRNA then recognizes a codon in mRNA, thereby delivering its amino acid to the growing polypeptide.aminoacyl-tRNAaminoacyl- tRNApolypeptide

Translation consists of three steps Initiation Elongation Termination

P siteA site PA mRNA binding site Each ribosome has a “P” and an “A” site. Codons mRNA Growing polypeptide tRNA Next amino acid to be added to polypeptide

- The process of decoding the information content of an mRNA into a linear sequence of linked amino acids is called translation. Translation requires the interaction of mRNA, charged tRNAs, ribosomes, and a large number of proteins (factors) that facilitate the initiation, elongation, and termination of the polypeptide chain. - In eukaryotic organisms, translation is initiated by the binding of a specific charged initiator tRNA, Met-tRNAMet, and other factors to the small ribosomal subunit. No other charged tRNA can bind to a free small ribosomal subunit. Next, the 5¢ end of an mRNA combines with the initiator tRNA–small ribosomal subunit complex, and the complex migrates along the mRNA until an AUG sequence (initiator codon) is encountered. Then, the UAC anticodon sequence of the initiator Met-tRNAMet base pairs with the AUG sequence of the mRNA, the migration stops, and the larger ribosomal subunit joins the complex. Initiation

1 Initiator tRNA mRNA Start codon Small ribosomal subunit 2 P site Large ribosomal subunit A site mRNA, a specific tRNA, and the ribosome subunits assemble during initiation Initiation

The elongation process continues until a UAA, UAG, or UGA codon is encountered. Elongation

1Codon recognition Amino acid Anticodon A site P site Polypeptide 2 Peptide bond formation 3 Translocation New peptide bond mRNA movement mRNA Stop codon

There are no naturally occurring tRNAs with anticodons that are complementary to UAA, UAG, or UGA (stop codons, termination codons). However, a protein (termination factor, release factor) recognizes a stop codon and binds to the ribosome. After binding of a termination factor, the bond between the last tRNA, which has the complete chain of amino acids linked to it, and its amino acid is broken. This cleavage results in the release of the uncharged tRNA, the complete protein, and the mRNA. Termination

- The AUG codon for methionine is the most common start codon. Synthesis of all protein chains in prokaryotic and eukaryotic cells begins with the amino acid methionine.codon startproteinamino acid - A reading frame, the uninterrupted sequence of codons in mRNA from a specific start codon to a stop codon, is translated into the linear sequence of amino acids in a protein.reading framestartcodonprotein - The three codons UAA, UGA, and UAG do not specify amino acids but constitute stop (terminator) signals that mark the carboxyl terminus of protein chains in almost all cells.

End Start of genetic message Translation begins at the start codon on the mRNA Initiation

Eukaryotic mRNA must be processed before it exits nucleus & enters cytoplasm. nucleotide cap is added “poly A tail” is added introns are removed

mRNA is in the language of nucleotides while polypeptides use the language of amino acids. To understand the information in mRNA, translation is needed! Where does translation take place?

The language of amino acids is based on codons 1 codon = 3 mRNA nucleotides 1 codon =1 amino acid A U A U A U G C C C G C

THE GENETIC CODE AND tRNA

Genetic code The set of rules whereby nucleotide triplets ( codons) in DNA or RNA specify amino acids in proteins. codon Sequence of three nucleotides in DNA or mRNA that specifies a particular amino acid during protein synthesis; also called triplet. Of the 64 possible codons, three are stop codons, which do not specify amino acids.

Using this chart, you can determine which amino acid the codon “codes” for! Which amino acid is encoded in the codon CAC?

Find the first letter of the codon CAC Find the second letter of the codon CAC Find the third letter of the codon CAC

CAC codes for the amino acid histidine (his).

What does the mRNA codon UAC code for? Tyr or tyrosine

Notice there is one start codon AUG. Transcription begins at that codon!

Notice there are three stop codons. Transcription stops when these codons are encountered.

Each amino acid is specified by more than one codon- degeneracy The Genetic Code

Degeneracy of the genetic code:Degeneracy: The genetic code is degenerate (sometimes called redundant). Although each codon corresponds to a single amino acid, a given amino acid may have more than one triplet coding for it. For example, arginine is specified by six different codons.

Degeneracy of the genetic code

THE CODE IS NEARLY UNIVERSAL Universality: The genetic code is virtually universal, that is, the specificity of the genetic code has been conserved from very early stages of evolution, with only slight differences in the manner in which the code is translated. [Note: An exception occurs in mitochondria, in which a few codons have meanings different than those shown in Nuclear DNA.( for example, UGA codes for trp.]

The results of large-scale sequencing of genomes have confirmed the universality of the genetic code. Benefits of the universal codes Make it possible to express cloned copies of genes encoding useful protein in different host organism. Example: Human insulin expression in bacteria)

However, in certain subcellular organelles, the genetic code is slightly different from the standard code.  Mitochondrial tRNAs are unusual in the way that they decode mitochondrial messages..

Genetic Code of Mammalian Mitochondria

What molecules are needed for translation? Ribosomes Transfer RNA (tRNA) mRNA Amino acids

All the amino acids that eventually appear in the finished protein must be present at the time of protein synthesis. [Note: If one amino acid is missing (for example, if the diet does not contain an essential amino acid), translation stops at the codon specifying that amino acid. This demonstrates the importance of having all the essential amino acids in sufficient quantities in the diet to ensure continued protein synthesis.

At least one specific type of tRNA is required per amino acid. In human, there are at least fifty species of tRNA, whereas bacteria contain thirty to forty species. Because there are only twenty different amino acids commonly carried by tANA, some amino acids have more than one specific tRNA molecule. This is particularly true those amino acids that are coded for by several codons. tRNA