SPIN - a new scaffold to design human peroxidase inhibitors

The human body has to defend itself constantly against invading pathogens such as viruses or bacteria. For this purpose, it has - among others - so called peroxidases, which are important parts of the innate immune system. Three peroxidases, called myeloperoxidase, eosinophil peroxidase and lactoperoxidase, are the focus of this study. Peroxidases are enzymes which produce small active molecules (radicals, oxidants) which are highly reactive and kill invading bacteria. These molecules, however, do not discriminate between bacteria and human cells. Therefore, if produced in the wrong place, e.g. in the blood or lung, they also attack the human tissue and lead to chronic inflammations. Therefore, a long-standing goal has been to identify substances, which can specifically block the production of radicals and to use them as inhibitors to treat and dampen chronic inflammations. Logically also bacteria have tried to find ways to survive and evade the immune system. Chief among them is Staphylococcus aureus, which has evolved a repertoire of proteins and other molecules to use as biological shields in its fight to escape. Among these, one small protein called SPIN (Staphylococcal peroxidase inhibitor) has been detected, which specifically targets myeloperoxidase. This protein, which is unlike any other known protein, has acquired the ability to specifically identify myeloperoxidase and uses a long flexible arm to block myeloperoxidase from producing radicals and oxidants. In this study the interaction between SPIN and myeloperoxidase will be studied in detail to understand the relationship between the structure of the two proteins and the inhibitory activity of SPIN. In addition, SPIN proteins from other Staphyloccocal species, which have different characteristics, will be studied. Together the knowledge will be used to design several different artificial specific SPIN inhibitors against the three peroxidases, which will have an increased binding strength and high specificity and will be better at blocking the undesired production of radicals and oxidants.

The human body has powerful defense systems to fight harmful bacteria and other microbes. Among these are the heme enyzmes, myeloperoxidase (MPO) and eosinophil peroxidase (EPO). Together with the related enzyme lactoperoxidase, they are essential parts of our innate immune system, the body's first line of defense. These enzymes produce highly reactive compounds that can kill or weaken invading pathogens in blood and in protective fluids covering our tissues. However, there is a downside: the same reactive substances that help fight infections can also damage the body's own tissues. This can trigger or worsen inflammation. When inflammation becomes chronic, as in atherosclerosis (the hardening of arteries), it can lead to serious health problems and place a growing burden on society, especially in aging populations. Because of this, scientists have long searched for drugs that can block the harmful effects of MPO and EPO without interfering with their useful roles. This has proven difficult, since MPO and EPO are very similar in structure to each other and to other enzymes of the same family. Blocking them in the wrong way could also disrupt other important processes, causing dangerous side effects. Designing safe and highly specific inhibitors therefore requires a deep understanding of the exact structural differences between these enzymes. Until recently, however, detailed structural data was only available for MPO. In this project, researchers achieved a major breakthrough by solving the first-ever crystal structure of native human EPO. This allowed, for the first time, a direct atomic-level comparison between MPO and EPO. They found subtle but important differences: variations in the electrical charge within the active site channel, differences in the flexibility of certain amino acids near the binding site, and unique features within the catalytic core. These differences explain why MPO and EPO behave differently when carrying out their chemical reactions. The study also investigated a small protein called SPIN, produced by the bacterium Staphylococcus aureus. SPIN has evolved to specifically block MPO, allowing the bacteria to escape part of the immune response. SPIN is surprisingly complex for its size: it has a compact globular domain that forms precise hydrogen bonds with MPO, and a flexible N-terminal loop that slowly inserts itself into MPO's active site. This insertion, driven by general hydrophobic effects, greatly reduces MPO's main activity-producing reactive chlorine compounds. The bi-partite nature of SPIN makes it extremely specific to MPO, but while the inhibition is strong it cannot be total. The insights gained in the project open the door to designing new drugs that could selectively block harmful peroxidase activity while leaving beneficial immune functions intact. This work lays the foundation for future laboratory research and clinical applications aimed at controlling chronic inflammation and its severe health consequences.