3R19

Chicken sulfite oxidase triple mutant with altered activity and substrate affinity

3R19 の概要

• エントリーDOI: 10.2210/pdb3r19/pdb
• 関連するPDBエントリー: 1sox 2A99 2BII 3HBG
• 分子名称: Sulfite oxidase, PHOSPHONIC ACIDMONO-(2-AMINO-5,6-DIMERCAPTO-4-OXO-3,7,8A,9,10,10A-HEXAHYDRO-4H-8-OXA-1,3,9,10-TETRAAZA-ANTHRACEN-7-YLMETHYL)ESTER, MOLYBDENUM ATOM, ... (4 entities in total)
• 機能のキーワード: molybdenum, molybdopterin, oxotransferase, sulfite oxidase, nitrate reductase, metal binding, nitrogen assimilation, oxidoreductase
• 由来する生物種: Gallus gallus (bantam,chickens)
• 細胞内の位置: Mitochondrion intermembrane space: P07850
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 51449.48

構造登録者

Qiu, J.A. (登録日: 2011-03-09, 公開日: 2012-02-08, 最終更新日: 2023-09-13)

主引用文献

Qiu, J.A.,Wilson, H.L.,Rajagopalan, K.V.
Structure-based alteration of substrate specificity and catalytic activity of sulfite oxidase from sulfite oxidation to nitrate reduction.
Biochemistry, 51:1134-1147, 2012

PubMed Abstract: Eukaryotic sulfite oxidase is a dimeric protein that contains the molybdenum cofactor and catalyzes the metabolically essential conversion of sulfite to sulfate as the terminal step in the metabolism of cysteine and methionine. Nitrate reductase is an evolutionarily related molybdoprotein in lower organisms that is essential for growth on nitrate. In this study, we describe human and chicken sulfite oxidase variants in which the active site has been modified to alter substrate specificity and activity from sulfite oxidation to nitrate reduction. On the basis of sequence alignments and the known crystal structure of chicken sulfite oxidase, two residues are conserved in nitrate reductases that align with residues in the active site of sulfite oxidase. On the basis of the crystal structure of yeast nitrate reductase, both positions were mutated in human sulfite oxidase and chicken sulfite oxidase. The resulting double-mutant variants demonstrated a marked decrease in sulfite oxidase activity but gained nitrate reductase activity. An additional methionine residue in the active site was proposed to be important in nitrate catalysis, and therefore, the triple variant was also produced. The nitrate reducing ability of the human sulfite oxidase triple mutant was nearly 3-fold greater than that of the double mutant. To obtain detailed structural data for the active site of these variants, we introduced the analogous mutations into chicken sulfite oxidase to perform crystallographic analysis. The crystal structures of the Mo domains of the double and triple mutants were determined to 2.4 and 2.1 Å resolution, respectively.

PubMed: 22263579
DOI: 10.1021/bi201206v

実験手法

X-RAY DIFFRACTION (2.1 Å)