24. Protein synthesis 1. Participants and mechanism

Last updated on November 19, 2018 at 17:16

Summary

  • The genetic code is the set of rules the ribosome uses to synthesize proteins from mRNA
    • = the codons of three bases and what amino acids they code for
  • The genetic code is degenerate, but unambiguous
    • This means that one amino acid can be coded by several codons (degenerate), but one codon codes for only one specific amino acid (unambiguous).
  • The genetic code is universal and conserved, which means it’s the same in nearly every species we know
  • Point mutations can be either silent, missense, nonsense or frameshift.
  • Some tRNAs, the molecules that carry the amino acids and bind to the correct codon on the mRNA, can recognize more than one codon by the function of wobble.
  • Aminoacyl-tRNA synthetase is the enzyme that binds the correct amino acid to tRNA. The enzyme needs ATP and Mg2+
  • The “second genetic code” is the interaction between Aminoacyl-tRNA synthetase and the tRNAs
  • Ribosomes are composed of protein and rRNA. The prokaryotic ribosome, 70S, is composed of 50S and 30S subunits. The eukaryotic ribosome, 80S, is much larger and is composed of 60S and 40S subunits.
  • Each tRNA has certain parts: a CCA sequence on the 3’-end which binds an amino acid, an anticodon arm, a TψC arm and a D arm. The anticodon on the anticodon arm is what recognizes the codon on the mRNA.
  • The polypeptide is synthetized in the direction from the amino-terminal to the carboxyl-terminal
  • Protein synthesis happens in five stages: amino acid activation, initiation, elongation, termination, and posttranslational processing and folding.

The genetic code

The genetic code. The green (AUG) is the start-codon, while the red are stop-codons. The codons are on the left while the three-letter codes of the corresponding amino acids are on the right.

The genetic code is basically a large table that shows which three RNA bases equals which amino acid in the finished protein. Three such RNA bases are called a codon.

Most amino acids are encoded by several codons (for example arginine is encoded by both AGA and AGG). This is what makes the genetic code degenerate. However, each codon only codes for 1 single amino acid. This is what makes the genetic code unambiguous. The codons on the mRNA are read in the 5’ -> 3’ direction.

The genetic code is the same (or very similar) in every species we know; it is universal and conserved (among species).

mRNA molecules are simply long chains of codons one after another. The ribosome reads one codon, connects the amino acid associated with that codon to the polypeptide, then reads the next codon and does the same, until it reaches a stop-codon.

2 types of point mutation. Note how the reading frame works.

Many types of mutations exist. Point mutations are mutations that either remove, add, or modify a base.

Some mutations are harmless. If the third base in the codon AUU is modified to a C, the polypeptide will be no different, because both AUU and AUC code for the same amino acid. This is called a silent mutation.

If the first base in CAA is modified to a U, the codon for glutamine suddenly becomes a stop codon. The ribosome will read this stop codon and stop the translation there, which will result in a half-finished polypeptide chain. This type of mutation is called nonsense mutation.

If the second base in GGA is modified to an A, the codon will code for glutamate instead of glycine. The ribosome will then insert a glutamate where there should have been a glycine in the polypeptide. This is called a missense mutation. Some missense mutations are not harmful, because many amino acids have similar properties, so switching between them don’t make for a big difference in the resulting protein.

Lastly, there is the frameshift mutation. If a new base is added or a present one is removed, the whole mRNA molecule becomes one base longer or shorter, respectively. This causes a shift in the reading frame. Consider the mRNA sequence GUAGCCUACGGA. If a C is inserted between the first UA, the mRNA sequence now becomes GUCAGCCUACGGA. The first mRNA sequence would be read like this: GUA – GCC – UAC – GGA, to yield the polypeptide Val – Ala – Tyr – Gly, but the second would be read like this: GUC – AGC – CUA – CGG – A, to yield the polypeptide Val – Ser – Leu – Arg. Notice how not only the mutated codon is modified, but every codon after it as well. This is because the reading frame was shifted one base to the right, which changes every single codon after it.

The ribosome and the tRNAs

The ribosome is an organelle that is composed of protein and rRNA. Ribosomes account for a lot of a cells weight. In E. coli there are 15000 ribosomes per cell, which account for 25% of the cells weight, water not included.

In both eukaryotes and prokaryotes, the ribosome is composed of two subunits. The prokaryotic ribosome is called 70S, and is composed of subunits 50S and 30S. The eukaryotic ribosome is called 80S, and is composed of subunits 60S and 40S.

The structure of tRNAs.

The tRNAs are the molecules that carry the amino acids to the ribosome, so they can be used as building blocks for the proteins. They bind to the different codons on the mRNA. There exists at least 32 different tRNAs. Each tRNA can bind 1-3 different codons, and can only carry one specific amino acid. The tRNA that can carry glycine is called tRNAGly, for example. When a tRNA has bound its corresponding amino acid, they called Met-tRNAMet, for example. If the tRNA that usually binds valine binds leucine instead, it would be a Leu-tRNAVal.

Each tRNA has a specific structure, shown on the picture on the last page. They have a D arm, an anticodon arm, a TψC arm, and an amino acid arm. The D arm is recognized by aminoacyl tRNA synthetase, which is the enzyme that binds the tRNA to the corresponding amino acid. The

TψC arm is bound by the ribosome. The amino acid arm binds the corresponding amino acid. The anticodon arm contains the anticodon, the part of the tRNA that recognizes the specific codon on the mRNA. The anticodon is composed of three bases, like a codon, but read in the 3’ -> 5’ direction, so opposite of the codon. For example, the anticodon on the tRNA that recognized the codon “GAA” is UUC.

As outlined above, some tRNAs can recognize more than one codon. This is because of “wobble”. The first nucleotide of the anticodon (which binds the third base on the codon), can be either C, A, U, G or I. The nucleotide I is derived from the base hypoxanthine, and can bind either U, C or A. If a tRNA has the anticodon IAC, it can bind to the following codons: GUU, GUC or GUA. Because the first nucleotide of the anticodon can bind multiple bases on the codon, we say that it “wobbles”.

This is how «wobbling» works. Some bases on the anticodon can bind to more than one base on the codon.
How one anticodon can recognize three different codons. We say that the anticodon “wobbles”.

 

 

Stage 1: Activation of amino acids

There exists one aminoacyl-tRNA synthetase for each amino acid-tRNA pair. During the first stage of protein synthesis, these enzymes “charge” the tRNAs with their corresponding amino acids. This reaction needs ATP and Mg2+. These enzymes proofread that they’ve bound the right amino acid to the right tRNA. The interaction between aminoacyl-tRNA synthetase and tRNAs is called the “second genetic code”.

The precise mechanism of protein synthesis is outside the scope of these notes, and probably the biochemistry exam as well.

Stage 2: Initiation

Protein synthesis begins at the amino-terminal end and proceeds by adding new amino acids to the carboxy-terminal end. The start codon, AUG, is also the only codon for methionine. Thus, all polypeptides synthesized begin with a methionine (N-formylmethionine in bacteria). The cell has two separate tRNAs for methionine, one that recognizes the AUG in the beginning of mRNAs, and one that recognizes the same codon is in the middle of an mRNA.

Proteins called initiation factors are needed for initiation. In eukaryotes, we need eIF3 (eukaryotic initiation factor 3), PolyA binding protein, eIF4B, eIF2, and finally eIF4F, which is a complex of eIF4E, eIF4G and eIF4A. eIF2 is the part that binds the Met-tRNA, the first amino acid of the polypeptide.

Stage 3: Elongation

Factors needed for elongation are EF-Tu, EF-Ts, EF-G, GTP, Mg2+ and aminoacyl-tRNAs. The formation of the peptide bonds themselves is catalysed by 23S rRNA, a ribozyme, a rRNA molecule with enzymatic activity.

Stage 4: Termination

In bacteria, three factors are needed, RF-1 to RF-3. However, eukaryotes use just one factor, eRF (eukaryotic releasing factor) for the same tasks. RF-1 and RF-2 recognize the three stop codons. They add the last amino acid to the chain, and then releases the chain. RF-3 dissociates the 70S ribosome into its 30S and 50S subunits. EF-G and IF-3 are also needed.

What’s the deal with selenocysteine?

Selenocysteine is an amino acid found in proteins, but there’s not codon for it. Instead, a serine is inserted at certain UGA codons, which is later selenylated to become selenocysteine.


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23. Retroviruses

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25. Protein synthesis 2. Regulation, posttranslational modifications

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