All organisms use cellular respiration to generate ATP – the energy currency of the cell. Part of this process is called ‘the respiratory chain’ where a series of proteins transport charged particles across a membrane. This results in the buildup of potential energy, similar to a battery, this energy is used to make ATP. In humans this process is coupled to the oxygen we breathe by using it in the final step of the respiratory chain. The protein that catalyzes this step is called the ‘terminal oxidase’. Bacteria have evolved to produce many different proteins that supplement the respiratory chain, this involves the production of terminal oxidases that use other molecules than oxygen. These alternative respiratory chains mean that these organisms are able to thrive in environments where oxygen is sparse.

I work on a protein involved in one such alternative respiratory chain in the bacterium Paracoccus denitrificans. This bacterium is part of the denitrification process in which nitrogen compounds from the soil are converted back into nitrogen gas. Paracoccus denitrificans can use nitrogen compounds in the ATP-making process, this allows it to live in environments with low oxygen. The terminal oxidase in this pathway is called cytochrome-c dependent Nitric Oxide Reductase (cNOR), this is the protein that I work on.

Nitric Oxide Reductase is similar to other terminal oxidases in that it has several heme cofactors which serve as electron-transporters through the protein. However, its active site contains iron where other terminal oxidases usually have a copper ion. Curiously the presence of this iron is also linked to the presence of two helper proteins, called chaperones. When we produce cNOR without these helper proteins, this iron is lost. We are trying to understand in detail how these helper proteins are involved in the insertion of the iron. The aim of my research is to understand this system and translate those findings to other similar systems as well. Eventually the knowledge of the chaperones can be used to aid in metal insertion processes when making artificial proteins, and from an environmental perspective it is interesting to understand the denitrification process as some of its intermediates are very potent greenhouse gases.