DNA Replication, Transcription & Translation

Whenever cells divide, they need to make an extra copy of their DNA through DNA replication. DNA gives our cells the ‘instructions’ to make proteins - the things which do the work in our cells and include enzymes, hormones and channel proteins. DNA is converted into protein in two processes which sound annoyingly similar - transcription and translation.

Semi-conservative DNA replication

During cell division, cells need to make a complete copy of their genetic information. When DNA is replicated, the new DNA molecule is made up of one strand of the original DNA whereas the other strand is made of freshly made DNA. Since half of the DNA is preserved from the previous round of DNA replication, we describe the process as semi-conservative. It takes place in the following stages:

• DNA helicase unwinds the double helix, breaking the hydrogen bonds between complementary base pairs to separate the strands. One of the strands will act as a template for synthesis of the other strand.

• Complementary nucleotides will attach to the template strand by hydrogen bonding.

• DNA polymerase catalyses the formation of phosphodiester bonds between nucleotides, forming a complementary strand alongside the template parent strand.

• Two daughter DNA molecules are formed, each containing half of the original DNA molecule.

It is important that DNA polymerase accurately copies the template strand to avoid placing the wrong DNA nucleotide in the incorrect position. To avoid this, DNA polymerase ‘proofreads’ the complementary strand as it moves along the DNA. If it detects a mismatch, it can ‘snip out’ the wrong nucleotide and replace it with the right one. DNA polymerase has an accuracy rate of about 99%, which means that mistakes do occur every once in a while. A mistake results in a change to the DNA base sequence, which is known as a mutation. DNA mutations can have detrimental effects to the organism, since an altered base sequence can change the sequence of amino acids in a protein, causing it to fold differently and possibly lose its function.

Meselson and Stahl’s Experiment

The evidence that DNA replication is semi-conservative comes from a pretty clever experiment carried out by Matthew Meselson and Franklin Stahl. Before this, scientists were unsure whether DNA replication was conservative or semi-conservative. If DNA replicated conservatively, the original DNA strands would remain intact and the newly synthesised DNA would consist of two freshly-made strands.

To figure this out they used a heavy isotope of nitrogen (N-15) which has an extra neutron compared to the normal, lighter form of nitrogen (N-14). They grew bacteria in the presence of the heavy nitrogen and any new DNA that the bacteria made would incorporate this isotope and so would weigh heavier. If DNA replicates conservatively, Meselson and Stahl knew that they’d see some DNA made of just heavy nitrogen with the rest made of only light nitrogen, but if it replicated semi-conservatively it would be a mixture of the two isotopes. Here’s how they carried out the experiment in more detail:

• Two populations of bacteria were grown - one in a solution containing heavy nitrogen and the other in a solution containing light nitrogen. As the bacteria grow and reproduce, they incorporate the nitrogen into their DNA.

• DNA was extracted from the bacteria and centrifuged to separate the DNA according to its weight. The DNA from the bacteria grown in N-15 separated at a higher density (a band lower down the test tube) compared to the DNA from the bacteria grown in N-14.

• The scientists then took the bacteria that had been growing in N-15 (which now just contains heavy DNA) and grew them in the light isotope N-14. Again, they extracted their DNA and separated it using centrifugation.

• After the first round of DNA replication, they saw just one band of DNA which was an intermediate weight between 14-N and 15-N. This indicates that the DNA was made up of both types of nitrogen isotopes (i.e. one old strand and one newly synthesised strand).

• After the second round of DNA replication, there was now 2 bands of DNA. One band has an intermediate weight but further newly synthesised DNA is now only being made using the lighter isotope. This proved that DNA is replicated semi-conservatively.

• If DNA replication is conservative, Meselson and Stahl would have seen two bands of DNA (one heavy and one light) after both the first and second rounds of replication.

Transcription

For a gene to produce a protein, the DNA within the gene must first be copied into RNA in a process called transcription. This is important because DNA is too big and wrapped snuggly around chromosomes to leave the nucleus. Just like photocopying a single useful page out of a chunky library book, the cell makes an RNA copy of all the important information contained in the gene. During transcription, RNA polymerase binds to the beginning of a gene in an area known as the promoter region. The promoter region is a regulatory region which does not code for amino acids but facilitates the process of transcription by helping RNA polymerase bind to the gene. RNA polymerase separates the DNA strands, producing a single DNA template for transcription. As RNA polymerase moves along one of the DNA strands (the template strand), it adds complementary nucleotides and connects them through the formation of phosphodiester bonds. The other strand is referred to as the coding strand and will have an identical sequence to the newly synthesised RNA, except for the presence of thymine instead of uracil. Eventually RNA polymerase will reach a codon which does not code for an amino acid but tells the enzyme to stop transcribing (these are called stop codons). A molecule of messenger RNA (mRNA) has been formed which will leave the nucleus and enter the cytoplasm.

Translation

Once in the cytoplasm, the messenger RNA finds its way to structures called ribosomes, which are basically protein-building machines. The ribosome attaches itself to the RNA and slides along it. The ribosome ‘reads’ the mRNA in a series of three bases (such as AUG, CCA, GCU) called codons. Each codon corresponds to a particular amino acid. As the ribosome reads the codons, a transfer RNA (tRNA) molecule which has a complementary anticodon carries an amino acid to the ribosome. Once the ribosome has read through the length of the mRNA, a series of different amino acids will have been dropped off by several tRNA molecules. Peptide bonds form between the amino acids to form a protein. tRNA molecules have an usual clover-shaped structure, formed by a single RNA strand folded over on itself through hydrogen bonding. At one end of the molecule, there is an amino-acid binding site and at the other there is an anticodon, which contains a complementary base sequence to the mRNA codon.

The Nature of the Genetic Code

The genetic code can be described in a number of ways - it is a triplet code, non-overlapping, degenerate and universal.

• Triplet code: three nucleotide bases make up a codon, which code for a particular amino acid.

• Non-overlapping code: the codons do not overlap. Once the ribosome has ‘read’ one codon and the appropriate amino acid has been recruited, the ribosome moves onto a new codon.

• Degenerate code: different codons can code for the same amino acid. For example, the codons CUU and CUC both code for the amino acid leucine. This means that some mutations will have no effect on the organism since the same protein will still be produced.

• Universal code: all organisms use the same genetic code. Bacteria, bonobos and bananas all contain DNA made up of the four nitrogenous bases that are found in humans.

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