Degradation of rutin by Aspergillus flavus. Purification of the dioxygenase, quercetinase

Abstract

MILLS. 1971. Degradation of rutin by Aspergillr~s pavrts. Purification of the dioxygenase, quercetinase. Can. J. Microbiol. 17: 11 1-1 18. Evidence is presented that a single enzyme, quercetinase, is responsible for the degradation of quercetin by Aspergillus flavlts to yield carbon monoxide and a depside, 2-protocatechuoylphloroglucinol carboxylic acid. A procedure for the isolation of the dioxygenase as a homogeneous protein is described. The most purified preparation degraded 10 800 umoles of quercetin/h mg protein and was homogeneous as judged by ultracentrifugation and by electrophoresis. The molecular weight was determined as 111 000 + 4000. K,,, values for quercetin and oxygen as substrates were 5.2 X 1 0 -W and 1.2 X 10-4 M respectively. The enzyme is a glycoprotein containing 27.5% carbohydrate and the amino acid composition is presented.

Anaerobic enzyme⋅substrate structures provide insight into the reaction mechanism of the copper-dependent quercetin 2,3-dioxygenase

“…This procedure reduced the residual glycosylation to Ϸ3-5% (wt͞wt). An activity assay, carried out according to the procedure of Oka et al (16), indicated that deglycosylation had no adverse effect on the catalytic efficiency of 2,3QD.…”

Copper Active Sites in Biology

“…Quercetinases have been isolated from the fungi Aspergillus flavus , 1099 Aspergillus niger , 1110 Aspergillus japonicus , 1111 and Penicillium olsonii 1112 and the prokaryotes Bacillus subtilis 1113,1114 and Streptomyces sp. FLA. 1115 All quercetinase enzymes are glycoproteins with 16–47% sugar content (w/w).…”

The Reaction of μ‐η2:η2‐Peroxo‐ and Bis(μ‐oxo)dicopper Complexes with Flavonol

“…Quercetinase has been reported to contain two Cu II centers per molecule. [19,20] Recent diffraction studies on single crystals of quercetin 2,3-dioxygenase obtained from Aspergillus japonicus, at a resolution of 1.8 Å , showed that the enzyme forms homodimers, which are stabilized by an Nlinked heptasaccharide at the dimer interface. [21] Each unit is mononuclear, with a type 2 copper center displaying two distinct geometries: a distorted tetrahedral coordination with three histidine units and one water molecules, and a distorted trigonal bipyramidal environment with an additional carboxylate.…”