Transcript

NARRATOR: Formation of the cisplatin DNA adduct has damaged the DNA. If the DNA is not repaired then the cell cannot reproduce itself or replicate. Research has moved on to examine the cell mechanisms that deal with this damage. Proteins are the key to processes such as the repair mechanism. Normally, damaged DNA is recognised and marked for repair, which is done by cutting out the damage, so-called excision repair.

The bent, platinated DNA seems to be recognised by other proteins which bind at the damaged site and prevent access by the repair proteins.

STEPHEN LIPPARD: The class of proteins that bind to the cisplatin-modified DNA, it turns out from gene sequencing and other experiments that we’ve done, generally contain what are called high-mobility group, or HMG, domains.

NARRATOR: The term ‘high-mobility group’ comes from this procedure known as electrophoresis. It’s used to identify and separate individual strands of DNA from a mix of all sorts. Cancer cells are cultured in the Petri dishes at body temperature, 37 degrees Celsius. The cells are harvested and the protein extracted.

They are then mixed with platinated DNA, which has also been made radioactive by including a radioactive isotope of phosphorus – 32 phosphorus – in the phosphate groups. The cultures are incubated for varying amounts of time and dyed a bright colour to make them visible.

The culture is dropped onto a gel and an electric field then applied. The DNA is negatively charged because of the phosphate groups. And so it begins to travel down the gel towards the positively charged cathode. To make a permanent record of the experiment, a photographic film is put next to the gel. The radioactive phosphorus isotope blackens the film, thus indicating the positions of the pieces of DNA.

As the repair mechanism operated during the incubation, bits of the DNA that are damaged by the cisplatin were excised. Because these pieces are smaller and lighter, they travel faster and further down the gel. The longer the incubation ran, the more small excised bits of platinum-containing DNA can be seen as the DNA tries to repair itself.

The technique can be used to study the effect of the HMG proteins on the repair mechanism.

STEPHEN LIPPARD: They could bind to the cisplatin-modified DNA and shield or protect the adduct from excision repair. If this happened more efficiently in a tumour cell than in a normal cell of the same tissue, that would be a wonderful mechanism for anticancer activity. And we’re working hard to evaluate that hypothesis.

NARRATOR: This structure shows an HMG protein – the red tube – attached to a natural binding site for transcription. It binds to the minor groove. But the similarity of the effect on the minor groove to the effect of cisplatin is striking. In both cases, the minor groove becomes wider and shallower. Perhaps it is possible that the cisplatin DNA confuses the repair mechanism by this similarity and so escapes excision.