The M42 aminopeptidases are dinuclear aminopeptidases displaying a peculiar tetrahedral-shaped structure with twelve subunits. Their quaternary structure results from the selfassembly of six dimers controlled by their divalent metal ion cofactors. The oligomeric state transition remains debated despite the structural characterization of several archaeal M42 aminopeptidases. The main bottleneck is the lack of dimer structures, hindering the understanding of structural changes occurring during the oligomerization process. We present the first dimer structure of an M42 aminopeptidase, TmPep1050 of Thermotoga maritima, along with the dodecamer structure. The comparison of both structures allows to describe how the metal ion cofactors modulate the active site fold and, subsequently, affect the interaction interface between dimers. A mutational study shows that the M1 site strictly controls dodecamer formation. The dodecamer structure of TmPep1050 also reveals that a part of the dimerization domain delimits the catalytic pocket and could participate in substrate binding.

The M42 aminopeptidases are a family of dinuclear aminopeptidases widely distributed in 16 Prokaryotes. They are potentially associated to the proteasome, achieving complete peptide 17 destruction. Their most peculiar characteristic is their quaternary structure, a tetrahedron-18 shaped particle made of twelve subunits. The catalytic site of M42 aminopeptidases is defined 19 by seven conserved residues. Five of them are involved in metal ion binding which is important 20 to maintain both the activity and the oligomeric state. The sixth conserved residue, a glutamate, 21 is the catalytic base deprotonating the water molecule during peptide bond hydrolysis. The 22 seventh residue is an aspartate whose function remains poorly understood. This aspartate 23 residue, however, must have a critical role as it is strictly conserved in all MH clan enzymes. 24 It forms some kind of catalytic triad with the histidine residue and the metal ion of the M2

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The M42 aminopeptidases are a family of dinuclear aminopeptidases widely distributed in Prokaryotes. They are potentially associated to the proteasome, achieving complete peptide destruction. Their most peculiar characteristic is their quaternary structure, a tetrahedron-shaped particle made of twelve subunits. The catalytic site of M42 aminopeptidases is defined by