STRUCTURE ANALYSIS OF B12 BINDING SUBUNIT OF GLUTAMATE MUTASE

依赖辅酶B12的谷氨酸变位酶催化 (S)-谷氨酸异构化为(2S，3S)-3-甲基天冬氨酸 四异形梭菌发酵谷氨酸的第一步。 为了更好地了解谷氨酸变位酶的机制，以及在 特别是载脂蛋白如何调节B12的反应性 辅因子，我们已经结晶了谷氨酸的B12结合亚基 变位酶(15 KDa)，辅酶B12(ADOCbl)存在时。晶体 通常获得1.2 mm×0.8 mm×0.7 mm的大小，以及 其中一些与2.2的分辨率不同。所有数据集都有 索引为具有单位晶胞尺寸a=b=的基元四方 104.7，c=162.7。较大的单元格表示 不对称单元可能包含四到六个分子。一本小说 利用ADOCbl的本征钴进行MAD定相的方法 一种反常的散射体，最近在国际象棋中被引导。两个数据 套装(91%完成至2.6%)所取的能量与 钴的峰值和拐点与以下数据相结合 一个家庭来源(1.5418辐射，用作远程波长)。四 在异常差值Patterson图上发现了钴的位置。 由Solve计算的相生成的电子密度图 无法解释。然而，分阶段的组合 钴同晶取代实验中的信息 从最近发现的汞衍生品中分阶段可能会证明 足以用于结构测定。

Coenzyme B12-dependent glutamate mutase catalyzes the isomerization of (S)-glutamate to (2S, 3S)-3-methylaspartate, the first step in glutamate fermentation by Clostridium tetanomorphum . To better understand the mechanism of glutamate mutase, and in particular how the apoprotein modulates the reactivity of the B12 cofactor, we have crystallized the B12-binding subunit of glutamate mutase (15 kDa), in the presence of coenzyme B12 (AdoCbl). Crystals as large as 1.2 mm x 0.8 mm x 0.7 mm have been obtained routinely, and some of them diffract to a resolution of 2.2
. All data sets have indexed as primitive tetragonal with unit cell dimensions a = b = 104.7
, c = 162.7
. The large unit cell indicates that the asymmetric unit likely contains four to six molecules. A novel approach to MAD phasing, employing the intrinsic cobalt of AdoCbl as an anomalous scatterer, was recently conducted at CHESS. Two data sets (91% complete to 2.6
) taken at energies corresponding to the peak and inflection points of cobalt were combined with data taken on a home source (1.5418
radiation, used as a remote wavelength). Four cobalt sites were identified in anomalous difference Patterson maps. Electron density maps generated from phases calculated by SOLVE have not been interpretable. However, the combination of phasing information from the cobalt experiments with isomorphous replacement phasing from a recently identified mercury derivative may prove adequate for structure determination.