Crystal Structure of Agmatinase Reveals Structural Conservation and Inhibition Mechanism of the Ureohydrolase Superfamily

Abstract

Agmatine is the product of arginine decarboxylation and can be hydrolyzed by agmatinase to putrescine, the precursor for biosynthesis of higher polyamines, spermidine, and spermine. Besides being an intermediate in polyamine metabolism, recent findings indicate that agmatine may play important regulatory roles in mammals. Agmatinase is a binuclear manganese metalloenzyme and belongs to the ureohydrolase superfamily that includes arginase, formiminoglutamase, and proclavaminate amidinohydrolase. Compared with a wealth of structural information available for arginases, no threedimensional structure of agmatinase has been reported. Agmatinase from Deinococcus radiodurans, a 304-residue protein, shows ϳ33% of sequence identity to human mitochondrial agmatinase. Here we report the crystal structure of D. radiodurans agmatinase in Mn 2؉ -free, Mn 2؉ -bound, and Mn 2؉ -inhibitor-bound forms, representing the first structure of agmatinase. It reveals the conservation as well as variation in folding, oligomerization, and the active site of the ureohydrolase superfamily. D. radiodurans agmatinase exists as a compact homohexamer of 32 symmetry. Its binuclear manganese cluster is highly similar but not identical to the clusters of arginase and proclavaminate amidinohydrolase. The structure of the inhibited complex reveals that inhibition by 1,6-diaminohexane arises from the displacement of the metal-bridging water.

Crystal structures of Pseudomonas aeruginosa guanidinobutyrase and guanidinopropionase, members of the ureohydrolase superfamily

“…GbuA and GpuA monomer structures are similar to other ureohydrolases (Ahn et al, 2004;Bewley et al, 1999;Cama et al, 2003;Di Costanzo et al, 2005;Elkins et al, 2002;Kanyo et al, 1996). According to the DALI search (Holm and Rosenström, 2010), S. clavuligerus PAH (PDB code 1GQ7; Elkins et al, 2002) shows the highest structural similarity with P. aeruginsa GbuA (chain A), (Thompson et al, 1997).…”

“…deviation of 1.9 Å for 299 equivalent C a positions, a Z-score of 35.3, and a sequence identity of 40%. The next highest Z-score is obtained with D. radiodurans agmatinase (PDB code 1WOG; Ahn et al, 2004), with an r.m.s. deviation of 2.2 Å for 291 equivalent C a positions, a Z-score of 33.0, and a sequence identity of 34%.…”

“…Previous reports showed that substrate recognition is mainly determined by the loops around the active site, which allows diverse guanidine substrates to be accommodated for catalysis (Ahn et al, 2004;Bewley et al, 1999;Cama et al, 2003;Di Costanzo et al, 2005Elkins et al, 2002;Kanyo et al, 1996;Shishova et al, 2009). Crystal structures of ureohydrolases are available for rat liver arginase I (Kanyo et al, 1996), human arginase I (Di Costanzo et al, 2005), human arginase II (Cama et al, 2003), Bacillus caldovelox arginase (Bewley et al, 1999), Streptomyces clavuligerus PAH (Elkins et al, 2002), and Deinococcus radiodurans agmatinase (Ahn et al, 2004). They contain a binuclear manganese cluster in the active site and exist as either a trimer or a hexamer.…”

“…aeruginosa GbuA and GpuA belong to the ureohydrolase superfamily, with 49% sequence identity between them. Other members of the ureohydrolase superfamily include arginase, agmatinase, proclavaminate amidinohydrolase (PAH), and formiminoglutamase, and they have narrow substrate specificities despite high levels of similarity in amino acid sequence and overall fold (Ahn et al, 2004;Bewley et al, 1999;Cama et al, 2003;Di Costanzo et al, 2005Elkins et al, 2002;Jann et al, 1988;Jenkinson et al, 1996;Kanyo et al, 1996;Itoh, 2002, 2005). Therefore, these enzymes may have evolved from a common ancestral protein and diverged into their particular functions with different substrate specificities (Arakawa et al, 2003;Dowling et al, 2008;Itoh, 2002, 2005;Sekowska et al, 2000).…”

Expression and localization of an agmatinase-like protein in the rat brain

“…However, expression of human agmatinase mRNA was reported to be most abundant in human liver and kidney, with a much lower expression in muscle, small intestine and brain (Iyer et al 2002). Human agmatinase has been cloned and its amino acid sequence showed a 35% identity to Escherichia coli agmatinase with all residues essential for metal binding and substrate hydrolysis by members of the ureohydrolases superfamily conserved, including all known agmatinases and arginases (Ahn et al 2004). The characterization of the enzyme was complicated by the extremely low catalytic activity of the recombinant protein (Iyer et al 2002;Mistry et al 2002).…”