Enzymatic metal-chelation by porphyrins

Sammanfattning: This thesis deals with the structure and function of two different enzymes, ferrochelatase and magnesium chelatase, that catalyse similar reactions. When this work started, a three-dimensional model of ferrochelatase existed. Much was known about the enzyme, but some details remained unclear. The focus has been to establish where the metal and the porphyrin binds to the enzyme, and how they are put together into one. From experiments performed, binding sites for the two substrates were established. It was found that ferrochelatase binds the porphyrin in a vice like grip and induces a distortion of the ring (paper II). The metal ion has been shown to bind close to the porphyrin (paper V). These findings taken together can provide a model for the catalytic mechanism of the enzyme (paper II and V). Magnesium chelatase, that consists of three different subunits, was a more open field. With access to a collection of barley mutants, deficient in chlorophyll biosynthesis, some questions regarding magnesium chelatase were intended to be answered. Three barley mutants, that each segregated in three distinguished phenotypes served as a starting point. These mutants were found to be deficient in the smallest subunit (paper I). To further characterise the mutants and how subunits interact, the corresponding mutations were introduced in a bacterial magnesium chelatase (paper IV). A big leap forward came with the three-dimensional structure of the smallest subunit (paper III) While the data presented in this thesis is converging future research on ferrochelatase to fewer questions, it has done quite the opposite for magnesium chelatase. Thanks to what is presented here, it is now known that the smallest magnesium chelatase subunit belongs to the AAA+ class of ATPases (paper III). Further it has been suggested that the middle sized subunit shows similarities to integrins (paper III), a group of cell surface receptors. The connection between AAA+ proteins and integrins was not previously mentioned, but later similar arrangements have been found in other proteins. However the mechanism of magnesium chelatase is still unknown. Hopefully the new aspects presented here will contribute to reveal its secrets.