醛缩酶A
硫矿硫化叶菌
果糖二磷酸醛缩酶
生物化学
醛缩酶B
酶
生物
化学
立体化学
古细菌
基因
作者
V. N. Zaĭtsev,Ulrike Johnsen,Matthias Reher,Marius Ortjohann,G.L. Taylor,Michael J. Danson,Peter Schönheit,S.J. Crennell
出处
期刊:Biochemistry
[American Chemical Society]
日期:2018-05-29
卷期号:57 (26): 3797-3806
被引量:4
标识
DOI:10.1021/acs.biochem.8b00535
摘要
The thermoacidophilic archaea Picrophilus torridus and Sulfolobus solfataricus catabolize glucose via a nonphosphorylative Entner–Doudoroff pathway and a branched Entner–Doudoroff pathway, respectively. Key enzymes for these Entner–Doudoroff pathways are the aldolases, 2-keto-3-deoxygluconate aldolase (KDG-aldolase) and 2-keto-3-deoxy-6-phosphogluconate aldolase [KD(P)G-aldolase]. KDG-aldolase from P. torridus (Pt-KDG-aldolase) is highly specific for the nonphosphorylated substrate, 2-keto-3-deoxygluconate (KDG), whereas KD(P)G-aldolase from S. solfataricus [Ss-KD(P)G-aldolase] is an enzyme that catalyzes the cleavage of both KDG and 2-keto-3-deoxy-6-phosphogluconate (KDPG), with a preference for KDPG. The structural basis for the high specificity of Pt-KDG-aldolase for KDG as compared to the more promiscuous Ss-KD(P)G-aldolase has not been analyzed before. In this work, we report the elucidation of the structure of Ss-KD(P)G-aldolase in complex with KDPG at 2.35 Å and that of KDG-aldolase from P. torridus at 2.50 Å resolution. By superimposition of the active sites of the two enzymes, and subsequent site-directed mutagenesis studies, a network of four amino acids, namely, Arg106, Tyr132, Arg237, and Ser241, was identified in Ss-KD(P)G-aldolase that interact with the negatively charged phosphate group of KDPG, thereby increasing the affinity of the enzyme for KDPG. This KDPG-binding network is absent in Pt-KDG-aldolase, which explains the low catalytic efficiency of KDPG cleavage.
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