Looking Inside The Cell: 3D-modelling a Key Molecule Involved in Cancer Development

The research teams in the ICR’s Section of Structural Biology apply techniques of X-ray crystallography, electron microscopy, biophysics, biochemistry and molecular biology to understand the physical characteristics of proteins and other molecules in the body that are involved in cancer development.

They produce a three dimensional ‘sculpture’ of the molecule of interest that not only allows scientists to see the shape and form of a molecule, but to investigate how the molecule is altered when it interacts with other proteins and how errors in the DNA code can change its structure. They can even model how drugs used to treat cancer can block or enhance the molecule’s behaviour.
Professor David Barford FRS  leads a team within the Section of Structural Biology that have recently published work in the prestigious scientific journal Nature* examining the structure of a molecule known as the anaphase-promoting complex (APC/C).  

Professor Barford says: “This molecule plays an important role in controlling cell division, and we know that it can be involved in the development of some cancers such as breast and colon. Understanding how this protein complex works will help us to understand its role in tumour development and to design new drugs that target this complex.”

APC/C is a very large molecule created by joining together 13 different proteins into a kind of molecular machine involved in regulating the cycle cell - a series of complex events that lead to the division, duplication (replication) and death of a cell. In cancer, the cell cycle is disrupted, allowing cells to multiply out of control.  

Specifically, the APC/C initiates the degradation of proteins called cyclins and securins by a process called ubiquitin-mediated proteolysis. Regulating the levels of cyclins and securins guides the cell through the processes of chromosome division and cell division, leading to the generation of two new daughter cells. In order for the events of chromosome and cell division to occur in an ordered sequence, the APC/C must regulate the degradation of these proteins at the appropriate time of the cell cycle.

To destroy proteins, APC/C must first find the proteins that need to be broken down and it does this by recognising ‘tags’ on proteins called destruction box (D-box) or KEN box domains. Scientists already knew that the process requires a co-activator protein (either a protein called Cdc20 or one called Cdh1) that combines with APC/C to recognise and lock onto the protein for degradation. However, how the 13 different components of APC/C fit together, how exactly the co-activator works with APC/C and how the complex recognises and then degrades targeted proteins has not been clear.

To find out more about this important molecule, Professor Barford’s team used electron microscopy and molecular biology, and in collaboration with Dr. Mark Williams at Birkbeck, University of London, nuclear magnetic resonance (NMR) spectroscopy, to determine the shape of the APC/C complex and created a 3D model to study how the complex works.  

Using electron microscopy, the team showed that most of the 13 subunits of APC/C form a scaffold or framework defining the complex’s triangular shape, but others form the key ‘sockets’ for the docking of substrate molecules like cyclins and securins. They saw that the co-activator Cdh1 fitted within the central cavity and was close to, but not bound, to the subunit Apc10. When D-box tagged proteins bound to the APC/C complex, the conformation of the complex changed so that Cdh1 and Apc10 formed a physical inter-connection, generating a receptor or ‘dock’ for D-box tagged proteins. Once bound, the protein could then be degraded.

The work by Professor Barford’s team has added to our understanding of how APC/C controls the cell cycle by targeting and destroying proteins, and this has important implications for cancer drug research. Prof Barford says: “Our studies on the APC/C are aimed at understanding how this large molecule works to control the cell cycle. Using a number of different techniques such as electron microscopy, protein crystallography and molecular biology, we have been able to develop a high-resolution 3D-picture of this complex.  This information will allow us to begin developing therapies that control APC/C activity in cancer”.

* Structures of APC/C(Cdh1) with substrates identify Cdh1 and Apc10 as the D-box co-receptor was published online in Nature on 24 November 2010.