Altering the binuclear manganese cluster of arginase diminishes thermostability and catalytic function.
Arginase is a thermostable (Tm = 75 degrees C) binuclear manganese metalloenzyme which hydrolyzes l-arginine to form l-ornithine and urea. The three-dimensional structures of native metal-depleted arginase, metal-loaded H101N arginase, and metal-depleted H101N arginase have been determined by X-ray crystallographic methods to probe the roles of the manganese ion in site A (Mn2+A) and its ligand H101 in catalysis and thermostability. We correlate these structures with thermal stability and catalytic activity measurements reported here and elsewhere [Cavalli, R. C., Burke, C. J., Kawamoto, S., Soprano, D. R., and Ash, D. E. (1994) Biochemistry 33, 10652-10657]. We conclude that the substitution of a wild-type histidine ligand to Mn2+A compromises metal binding, which in turn compromises protein thermostability and catalytic function. Therefore, a fully occupied binuclear manganese metal cluster is required for optimal catalysis and thermostability.
Scolnick
L R
LR
Department of Chemistry, University of Pennsylvania, 231 South 34th Street, Philadelphia, Pennsylvania 19104-6323, USA.
Kanyo
Z F
ZF
Cavalli
R C
RC
Ash
D E
DE
Christianson
D W
DW
D000818:Animals; D001119:Arginase / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002384:Catalysis; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D004789:Enzyme Activation; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D008099:Liver / Q000201:enzymology; D008345:Manganese / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D008667:Metalloproteins / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D015394:Molecular Structure; D011487:Protein Conformation; D017510:Protein Folding; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013696:Temperature
D008667:Metalloproteins; D011994:Recombinant Proteins; D008345:Manganese; D001119:Arginase