Insight on the interaction of an agmatinase-like protein with Mn2 + activator ions

Abstract Agmatinase is an enzyme that catalyzes the hydrolysis of agmatine, a compound that is associated with numerous functions in the brain of mammalian organisms such as neurotransmitter, anticonvulsant, antinociceptive, anxiolytic and antidepressant-like actions. To date the only characterized agmatinases with significant enzymatic activity were extracted from bacterial organisms. These agmatinases are closely related to another ureahydrolase, arginase; both have binuclear Mn 2 + centers in their active sites. An agmatinase-like protein (ALP) from rat brain was identified that bears no sequence homology to known agmatinases (E. Uribe, M. Salas, S. Enriquez, M.S. Orellana, N. Carvajal, Arch. Biochem. Biophys. 461(2007) 146–150). Since all known ureahydrolases contain histidines in their binuclear Mn 2 + site each of the five histidine residues in ALP was individually replaced by alanines to identify those that may be involved in metal ion binding. Reactivation assays and thermal stability measurements indicated that His206 is likely to interact with a Mn 2 + bound to a high affinity site. In contrast, His65 and possibly His435 are important for binding of a second Mn 2 + to a lower affinity site. Metal ion binding to that site is not only leading to an increase in reactivity but also enzyme stability. Thus, similar to bacterial agmatinases and some of the antibiotic-degrading, Zn 2 + -dependent metallo-β-lactamases ALP appears to be active in the mono and binuclear form, with binding of the second metal ion increasing both reactivity and stability.