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PDBsum entry 1hqx
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Subunit-Subunit interactions in trimeric arginase. Generation of active monomers by mutation of a single amino acid.
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Authors
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L.T.Lavulo,
T.M.Sossong,
M.R.Brigham-Burke,
M.L.Doyle,
J.D.Cox,
D.W.Christianson,
D.E.Ash.
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Ref.
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J Biol Chem, 2001,
276,
14242-14248.
[DOI no: ]
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PubMed id
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Abstract
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The structure of the trimeric, manganese metalloenzyme, rat liver arginase, has
been previously determined at 2.1-A 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 S-shaped oligomerization motif at the carboxyl
terminus of the protein that mediates approximately 54% 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 degrees C, while the corresponding
value for the wild-type enzyme is 70 degrees C. The three-dimensional structure
of the R308K variant has been determined at 3-A 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.
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Figure 1.
Fig. 1. The crystal structure of rat liver arginase
showing the trimeric quarternary structure, S-shaped tail in
white at the subunit interface, and two manganese ions (spheres)
at the active site of each subunit. Arg-308 is represented in
the white ball and stick form.
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Figure 4.
Fig. 4. Differential scanning calorimetry data for the
unfolding of wild-type arginase (solid line), R308A (dashed and
dotted line), R308E (dotted line), and R308K (dashed line) shown
as excess molar heat capacity versus temperature. All scans are
normalized per mole of arginase subunit. Conditions were as
follows: 20 mM NaPO[4], 150 mM NaCl, 0.1 mM MnCl[2], pH 7.4, and
0.4 mg/ml arginase.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2001,
276,
14242-14248)
copyright 2001.
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