Interaction between the biotin carboxyl carrier domain and the biotin carboxylase domain in pyruvate carboxylase from Rhizobium etli.
Interaction between the biotin carboxyl carrier domain and the biotin carboxylase domain in pyruvate carboxylase from Rhizobium etli.
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生物素羧基载体结构域与丙酮酸羧化酶中的生物素羧化酶结构域之间的相互作用。
DOI：
10.1021/bi201277j
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发表时间：
2011-11-15
期刊：
影响因子：
2.9
通讯作者：
St Maurice, Martin
中科院分区：
文献类型：
--
作者：
Lietzan, Adam D.;Menefee, Ann L.;Zeczycki, Tonya N.;Kumar, Sudhanshu;Attwood, Paul V.;Wallace, John C.;Cleland, W. Wallace;St Maurice, Martin
丙酮酸羧化酶（PC）催化丙酮酸羧化为草酰乙酸，这是哺乳动物组织中重要的回补反应。为了实现催化作用，PC的拴系生物素必须能够进入生物素羧化酶结构域和羧基转移酶结构域中的活性位点。先前的研究已经证明，来自Rhizobium etli的PC中的苏氨酸882突变为丙氨酸使得羧基转移酶结构域失活，并且有利于生物素在生物素羧化酶结构域中的定位。我们报告的2.4 μ m分辨率的X-射线晶体结构的根瘤菌etli PC T882 A突变体，揭示了第一个高分辨率的描述域之间的相互作用的生物素羧基载体蛋白结构域和生物素羧化酶结构域。整体四元排列的根瘤菌etli PC保持高度不对称，是独立的变构激活剂的存在下。虽然在生物素羧化酶结构域中观察到生物素，但Arg 353和Glu 248之间的相互作用阻止了其进入活性位点，揭示了调节羧生物素进入BC结构域活性位点的机制。生物素羧基载体蛋白结构域的结合位置表明，拴系的生物素不能以与游离生物素相同的方向结合在生物素羧化酶结构域活性位点中，这有助于解释在生物素羧化酶中拴系的生物素和游离生物素底物之间观察到的催化差异。位于生物素羧化酶结构域活性位点的电子密度被分配给膦酰乙酸，提供了一个可能的位置，为推定的羧基磷酸中间体形成生物素羧化。从T882 A根瘤菌etli PC晶体结构中获得的见解提供了一系列新的PC催化快照，并对生物素依赖性酶家族的催化作用提供了一个新的视角。
Pyruvate carboxylase (PC) catalyzes the ATP-dependent carboxylation of pyruvate to oxaloacetate, an important anaplerotic reaction in mammalian tissues. To effect catalysis, the tethered biotin of PC must gain access to active sites in both the biotin carboxylase domain and the carboxyl transferase domain. Previous studies have demonstrated that a mutation of threonine 882 to alanine in PC from Rhizobium etli renders the carboxyl transferase domain inactive and favors the positioning of biotin in the biotin carboxylase domain. We report the 2.4 Å resolution X-ray crystal structure of the Rhizobium etli PC T882A mutant which reveals the first high-resolution description of the domain interaction between the biotin carboxyl carrier protein domain and the biotin carboxylase domain. The overall quaternary arrangement of Rhizobium etli PC remains highly asymmetrical and is independent of the presence of allosteric activator. While biotin is observed in the biotin carboxylase domain, its access to the active site is precluded by the interaction between Arg353 and Glu248, revealing a mechanism for regulating carboxybiotin access to the BC domain active site. The binding location for the biotin carboxyl carrier protein domain demonstrates that tethered biotin cannot bind in the biotin carboxylase domain active site in the same orientation as free biotin, helping to explain the difference in catalysis observed between tethered biotin and free biotin substrates in biotin carboxylase enzymes. Electron density located in the biotin carboxylase domain active site is assigned to phosphonoacetate, offering a probable location for the putative carboxyphosphate intermediate formed during biotin carboxylation. The insights gained from the T882A Rhizobium etli PC crystal structure provide a new series of catalytic snapshots in PC and offer a revised perspective on catalysis in the biotin-dependent enzyme family.
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DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
4.8
作者：
Hasan, Noaman M.;Longacre, Melissa J.;MacDonald, Michael J.
通讯作者：
MacDonald, Michael J.
影响因子：
37.3
作者：
Fan TW;Lane AN;Higashi RM;Farag MA;Gao H;Bousamra M;Miller DM
通讯作者：
Miller DM
DOI：
10.1073/pnas.77.3.1321
发表时间：
1980-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
FREYTAG, SO;UTTER, MF
通讯作者：
UTTER, MF
影响因子：
4.8
作者：
Janiyani, K;Bordelon, T;Cronan, JE
通讯作者：
Cronan, JE