Subunit-Subunit Interactions in Trimeric Arginase

Abstract

The structure of the trimeric, manganese metalloenzyme, rat liver arginase, has been previously determined at 2.1-Å resolution (Kanyo, Z. F., Scolnick, L. R., Ash, D. E., and Christianson, D. W., (1996) Nature 383, 554 -557). A key feature of this structure is a novel Sshaped oligomerization motif at the carboxyl terminus of the protein that mediates ϳ54% of the intermonomer contacts. Arg-308, located within this oligomerization motif, nucleates a series of intramonomer and intermonomer salt links. In contrast to the trimeric wild-type enzyme, the R308A, R308E, and R308K variants of arginase exist as monomeric species, as determined by gel filtration and analytical ultracentrifugation, indicating that mutation of Arg-308 shifts the equilibrium for trimer dissociation by at least a factor of 10 5 . These monomeric arginase variants are catalytically active, with k cat /K m values that are 13-17% of the value for wild-type enzyme. The arginase variants are characterized by decreased temperature stability relative to the wild-type enzyme. Differential scanning calorimetry shows that the midpoint temperature for unfolding of the Arg-308 variants is in the range of 63.6 -65.5°C, while the corresponding value for the wild-type enzyme is 70°C. The three-dimensional structure of the R308K variant has been determined at 3-Å resolution. At the high protein concentrations utilized in the crystallizations, this variant exists as a trimer, but weakened salt link interactions are observed for Lys-308.

Adaptations to a Subterranean Environment and Longevity Revealed by the Analysis of Mole Rat Genomes

“…His254 is located immediately downstream of a conserved motif required for binding manganese and ARG1 function (Dowling et al, 2008) (Figure 2C). Moreover, ARG1 is a homotrimer, with the salt bridges formed by Arg255 and Glu256 critical for its assembly (Lavulo et al, 2001; Sabio et al, 2001). The charged residue flanking the ARG1 core may improve ammonia removal efficiency by interacting with the acidic Glu256 or by strengthening the Arg255-Glu256 salt bridge.…”

Crystal Structure of Schistosoma mansoni Arginase, a Potential Drug Target for the Treatment of Schistosomiasis

“…The S-shaped C-terminus is suggested to be important for oligomerization and mediates 54% of the intermonomer contact surface area in rat arginase I, 49 although mutagenesis studies suggest that mutations in the C-terminus destabilize but do not necessarily prevent trimerization of rat arginase I or human arginase I. 50–52 The unusually long C-terminal tail of SmARG is similarly responsible for the majority of subunit-subunit interactions; the 13-residue extension alone contributes ~3,900 Å 2 total buried surface area (36% of total buried surface area) to trimer assembly.…”

Homology modeling, docking and molecular dynamics of the Leishmania mexicana arginase: A description of the catalytic site useful for drug design

“…In the rat liver arginase, the Arg308 residue forms an intramonomer salt bridge with the Glu262 and an intermonomer salt bridge with the Asp204 on the adjacent subunit [50]. Both Glu262 and Asp204 residues are conserved in the arginases of rats, humans, mice and several species of frogs of the genus Xenopus.…”