RiboNucleic Acids
Genetic Information is expressed through Nucleic Acids chains called RiboNucleic Acids.
They are three types of RNAs differing in size, function and localization:
Messenger RNA (mRNA) is a carrier of genetic information, a copy of a gene sequence acting as a template for protein construction.
Ribosomal RNA (rRNA) and Transfer RNA (tRNA) (also sometimes referred to as insoluble and soluble RNAs) are structural ribonucleic acids wich support the expression of mRNA into protein.
Messenger RNA (mRNA) is a carrier of genetic information, a copy of a gene sequence acting as a template for protein construction.
Ribosomal RNA (rRNA) and Transfer RNA (tRNA) (also sometimes referred to as insoluble and soluble RNAs) are structural ribonucleic acids wich support the expression of mRNA into protein.
Structures
RNAs are polynucleotides chains wich differ from those of DNAs by having ribose sugar instead of deoxyribose and uracil bases(U) instead of thymines (T).
double The hydroxyl function in 2' of ribose greatly affects the properties of RNAs. In particularly this enables more tertiary interactions wich tend to destabilize the 5'-3' phosphodiester bonds and prevent RNAs from adopting a B helix conformation.
However RNAs are single stranded molecules that often fold on themselves by bases pairing, thus forming structures called hairpin loops. Thus, excepting mRNAs wich display smooth linear structure, tRNAs and rRNAs adopt specified tertiary structures in association with proteins.
Transcription
The process in wich DNA is converted into a complementary RNA (RiboNucleic Acid) strands is called Transcription. It involves a powerfull enzymatic complex called RNA polymerase holoenzyme. This enzyme unravels and unzips DNA helix, recruits RNA nucleotides and matches them by base pairing to the DNA gene sequence.
The process in wich DNA is converted into a complementary RNA (RiboNucleic Acid) strands is called Transcription. It involves a powerfull enzymatic complex called RNA polymerase holoenzyme. This enzyme unravels and unzips DNA helix, recruits RNA nucleotides and matches them by base pairing to the DNA gene sequence.
The transcription is rather similar in prokaryotes and eukaryotes. One of the differences is that eukariotic cells possess three different types of RNA polymerases (I, II, III), instead of one in prokaryotes. Each type of eukaryotic RNA polymerase is responsible for the synthesis of a class of RNAs (pol I for rRNAs, pol II for mRNAs and pol III for tRNAs and 5S rRNAs).
Transcription is classicaly described in three distinct steps: initiation, elongation and termination.
Initiation occurs when the RNA polymerase holoenzyme binds at a special sequence in DNA called a promoter. The promoter consists of consensus sequences containing specific strings like TATA (Pribnow box) and CAAT (in eukaryotes).
An additional small protein, the factor sigma, attaches to the polymerase and stabilises it, locking it on the DNA strand to be transcripted. Then, the polymerase separates the double stranded DNA to form a bubble allowing the first nucleoside triphosphate to pair with the complementary DNA nucleotide .
An additional small protein, the factor sigma, attaches to the polymerase and stabilises it, locking it on the DNA strand to be transcripted. Then, the polymerase separates the double stranded DNA to form a bubble allowing the first nucleoside triphosphate to pair with the complementary DNA nucleotide .
Elongation of the RNA chain involves successive addition of nucleotides in the 5' to 3' direction.
Termination occurs when a Stop signals indicating the end of the gene is encountered. The termination signal is generaly a GC-rich palindrome forming a local stem-loop structure in the RNA , followed by an oligo A region. This sequence disrupts the base pairing of newly synthesized RNA with the DNA template, forcing the RNA and the polymerase to fall off. Sometimes termination also involves a specific protein (Rho protein).
In prokaryotes cells, transcription takes place in cytoplasm. When transcription is completed, RNAs are immediatly ready for use in translation. Translation can even begin during transcription thus allowing typical regulations process.
In prokaryotes cells, transcription takes place in cytoplasm. When transcription is completed, RNAs are immediatly ready for use in translation. Translation can even begin during transcription thus allowing typical regulations process.
In contrast, eukaryotic transcription takes place in nucleus. The RNAs primary transcripts, sometimes called Heterogenous Nuclear RNA (hnRNA) are often modified in the nucleus before export to the cytoplasm.
In particular, eukaryotic mRNAs undergo extensive modifications to increase their stability and become biologically active.
Thus, the 5' end of mRNAs is capping with a 7-methylguanosine (7mGTP) shortly after initiation. The unique 5' - 5' triphosphate linkage formed increase mRNA stability by affording protection from exonucleases. It also brings a recognizable signal for proteins involved in subsequent splicing process and also during translation.
Messenger RNAs are also polyadenylated at the 3' end. Just before termination a specific sequence, AAUAAA, is recognized by a polyadenylate polymerase. The primary transcript is cleaved approximately 20 bases downstream and a string of 20 - 250 Adenines termed poly-A tail is added to the 3' end.
Since a primary transcript is a mirror copy (negatif) of all the gene sequence it includes also intronic non-conding sequences. Therefore, a post-transcriptional modification of major importance consist in introns removal in a process called RNA splicing.
The mechanism involves formation of a loop, called a lariat, in a process directed by small nuclear ribonucleoproteins (snRNPs). The complex mRNA-snRNPs is called a spliceosome.
Some proteins are often attached to exported mRNAs forming ribonucleoprotein particles (mRNP). These mRNPs are supposed to help in transport through the nuclear pores and also in binding to ribosomes.
Translation
Proteins are the major structural and functional constituants of the cells. A protein exhibit a complex molecular structure formed by polypeptide chains made of basic subunits called Amino Acids.
Expression of a mRNA code into a polypeptide chain is named Translation. This complex process requires all three classes of RNAs.
mRNAs
In the mRNA code, each amino acid is designated by a triplet of nucleotides called codon. The genetic code consist of 64 different codons (4 bases: 4x4x4 possibilities for a triplet). Three triplets are Stop codons (termination codons) which stop the process of translation. The remaining 61 codons encode 20 different amino acids. Since several codons encode a same aminoacid the genetic code is thus degenerated (or redundant).
Proteins are the major structural and functional constituants of the cells. A protein exhibit a complex molecular structure formed by polypeptide chains made of basic subunits called Amino Acids.
Expression of a mRNA code into a polypeptide chain is named Translation. This complex process requires all three classes of RNAs.
mRNAs
In the mRNA code, each amino acid is designated by a triplet of nucleotides called codon. The genetic code consist of 64 different codons (4 bases: 4x4x4 possibilities for a triplet). Three triplets are Stop codons (termination codons) which stop the process of translation. The remaining 61 codons encode 20 different amino acids. Since several codons encode a same aminoacid the genetic code is thus degenerated (or redundant).
tRNAs
Transfert RNAs (tRNA) act as adapter between nucleotides codons and amino acids. They pick up free amino acids in cytoplasm and carry them into the ribosomes where polypeptide chain is elongated.
Transfert RNAs (tRNA) act as adapter between nucleotides codons and amino acids. They pick up free amino acids in cytoplasm and carry them into the ribosomes where polypeptide chain is elongated.
tRNAs are polynucleotide of about 60 - 95 nucleotides long, including few specific nucleotids (dihydro-uridine, pseudo-uridine).
They exhibit a cloverleaf-like secondary structure consisting of a stem and three main loops. They also display a tertiary L-like structure, which interacts with ribosomes.
They exhibit a cloverleaf-like secondary structure consisting of a stem and three main loops. They also display a tertiary L-like structure, which interacts with ribosomes.
The larger loop include a specific nucleotide triplet, the anticodon, wich may bind to a complementary codon of a mRNA.
The stem ends in 3' by the sequence ...CCA, which is the attachment site for an amino acid Each tRNA is coupled to the amino acid in accordance with its anticodon. The coupling between a given tRNA and the corresponding amino acid is catalyzed by a specific aminoacyl-tRNA synthetases.
The different tRNAs that accept a given amino acid are called isoacceptors.
Obviously, there should be as many different tRNAs as meaning codons (ie 61). In fact there is generally at most 56 different type of tRNAs in any cell. Therefore it seems that some tRNAs are able to recognize at least two of the different codons specifying a given amino acids (Wobble hypothesis).
Obviously, there should be as many different tRNAs as meaning codons (ie 61). In fact there is generally at most 56 different type of tRNAs in any cell. Therefore it seems that some tRNAs are able to recognize at least two of the different codons specifying a given amino acids (Wobble hypothesis).
rRNAs
The rRNAs are the major constituents of ribosomes.
Ribosomes are the cell organelles where the mRNA is read and translated into a protein sequence. A Ribosome holds the mRNA in place, matches the anti-codon of a tRNA carring appropriate amino acid, to the complementary codon of the mRNA and catalyses the peptide bonds formation.
The rRNAs are the major constituents of ribosomes.
Ribosomes are the cell organelles where the mRNA is read and translated into a protein sequence. A Ribosome holds the mRNA in place, matches the anti-codon of a tRNA carring appropriate amino acid, to the complementary codon of the mRNA and catalyses the peptide bonds formation.
A ribosome consists of two subunits of different size containing rRNAs arranged with specific proteins. Both rRNAs and associated proteins are slightly different in prokaryotes vs eukaryotes.
- The larger subunit (50S/60S) countains two rRNA molecules (5S + 23S / 5S + 28S) (S sedimentation coefficient, measures the relative size). It displays two binding sites for tRNAs : the peptidyl-tRNA (P) site and the aminoacyl-tRNA (A) site.
- The smaller subunit (30s/40S) which is made of one rRNA molecule
(16S / 18S) possesses a binding site for the mRNA.
Translation steps
Translation proceeds in cytoplasm in an ordered process. It requires free amino acids, free energy, mRNA, tRNAs, Ribosomes, and several non-ribosomal protein factors (eIF in Eukaryotes and IF in some prokaryotes).
The first phase called Initiation begins with the formation of an preinitiation complex between the small ribosomal unit, a protein factor (eIF2 or IF2) and an initiator tRNA carrying a methionine (tRNAmeti).
When the complex encounter a mRNA it recognize a specific sequence (Shine-Delgarno for prokaryotes or 5'Cap for eukaryotes) and pair the initiator codon AUG to the initiator tRNA anticodon (UAG).Then the larger ribosomal subunit associates with the initiation complex, thus matching the initiator tRNA at P site.
A next tRNA carrying an other amino acid is attracted and pairs with the next codon at the A site, the first peptide bond is catalysed by a ribosomal protein (peptidyl-transferase).
When the complex encounter a mRNA it recognize a specific sequence (Shine-Delgarno for prokaryotes or 5'Cap for eukaryotes) and pair the initiator codon AUG to the initiator tRNA anticodon (UAG).Then the larger ribosomal subunit associates with the initiation complex, thus matching the initiator tRNA at P site.
A next tRNA carrying an other amino acid is attracted and pairs with the next codon at the A site, the first peptide bond is catalysed by a ribosomal protein (peptidyl-transferase).
During the second phase named Elongation the ribosome continues to read codons from the 5' to the 3' and amino acids are added to the C-terminal growing peptide.
During each peptide bond formation, the polypeptide attached to the tRNA in the P site is transferred to the amino group of the aminoacyl-tRNA in the A site (Transpeptidation). Then the ribosome moves to the next codon. The empty tRNA is ejected and the peptidyl-tRNA is shifted from the A site to the P site (Translocation). A new aminoacyl-tRNA is allowed to enter within the A site.
During each peptide bond formation, the polypeptide attached to the tRNA in the P site is transferred to the amino group of the aminoacyl-tRNA in the A site (Transpeptidation). Then the ribosome moves to the next codon. The empty tRNA is ejected and the peptidyl-tRNA is shifted from the A site to the P site (Translocation). A new aminoacyl-tRNA is allowed to enter within the A site.
Termination phase arrives when a stop codon is reached. Stop codons are triplets which are not recognized by any tRNA (UAA, UAG, UGA), but by a protein releasing factor (RF1 or RF2 in prokaryotes, eRF in eukaryotes). The factor R binds to the A site and causes the release of the polypeptide chain. The inactive ribosome then releases the mRNA and dissociates its sub-units.
It should be noted that the polypeptide sequence is in total agreement with the gene code since tRNA anticodons are complementary of mRNA codons and the mRNA sequence is a mirror of the gene DNA sequence.
Several ribosomes can progress along the same mRNA strand, each making one polypeptide chain. These clusters called polysomes are free in the cytoplasm or may be binded to particular cell organelles that store proteins (rough endoplasmic reticulum).
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