Structural characterization of proteins to investigate their roles in diseases: Focus on MID & LTA4H

Abstract: Protein molecules are responsible for many biological functions in cells. In order to fulfill their various biological roles, these chain-like molecules must fold into precise three-dimensional shapes. The knowledge of accurate molecular structures is a prerequisite for rational drug design and for structure based functional studies. Getting structural information of proteins can be a very difficult task, especially when it comes to high resolution. Moraxella catarrhalis is widely recognized human-restricted gram-negative bacterium for which it has become clear that it is a true pathogen of both the upper and lower respiratory tract. After Haemophilus influenzae and Streptococcus pneumonia, it is the third most common cause of otitis media in children. The bacterium can directly stimulate B-cells without any recognition of T-cells and it can therefore be classified as a T-cell independent antigen. The mitogenic activity of Moraxella catarhallis is performed by a 2139 residue long outer membrane protein MID. An IgD binding domain (MID962-1200) has been described and the colonization to human respiratory tract cells is mediated by a 150-residue adhesin domain (MID764-913). SAXS studies on the IgD binding domain showed that this domain has an elongated 3-fold organization and that there is the presence of unordered/flexible structures. CD data and prediction of secondary structure for both of the domains indicated the presence of large amounts of (∼33%) ß-sheet and ∼10% α-helix content. Native datasets for MID962-1200 to 2.3 Å resolution and for MID764-913 to 2.7 Å resolution are collected and processed. Vertebrate leukotriene A4 hydrolases are zinc metalloenzymes with an epoxide hydrolase and aminopeptidase activity belonging to the M1 family of aminopeptidases. The human enzyme produces LTB4, a powerful mediator of inflammation and is implicated in a wide variety of rheumatoid diseases. The yeast homolog scLTA4H contains only a rudimentary epoxide hydrolase activity and was shown to undergo a large conformational change upon binding of the inhibitor bestatin. In SAXS studies XlLTA4H shows a more compact form upon bestatin binding, but humLTA4H did not. It was confirmed that the LTA4H from Xenopus is a dimer and that it seems to contract in size upon bestatin binding. In contrast the human enzyme does not show any major difference in SAX scattering patterns upon inhibitor binding and it seems therefore that this enzyme does not display larger conformational changes.