Structural Analysis of Mutants of the Mouse Major Urinary Protein

The mouse Major Urinary Proteins (MUPs), known also as Mus m 1 urinary allergen complex , are lipocalins involved in pheromonal signalling. The 3D structure of a recombinant member of this protein family, rMUP, has been determined by NMR spectroscopy and crystallography. The lipocalin fold creates an internal cavity recognized as the binding site for a range of hydrophobic ligands acting as chemosignals perceived by conspecifics. It was previously suggested that the binding of ligands inside the pocket relies upon the formation of a network of interactions with selected protein side chains. The aim of this work is to elucidate the functional and structural role of Tyr120 whose side chain appears to interact with the bound pheromone 2-sec-buthyl-thyazoline. By site-specific mutagenesis this residue was replaced with Phe (rMUP-Y120F) and Ala (rMUPY120A). Fluorescence and CD spectroscopy data taken together with guanidinium chloride denaturation results show that Tyr120 not only plays an important role in the binding process but also is a key residue in the stabilization of the protein architecture.