Department of Biochemistry and Food Chemistry, University of Turku, Finland.
Biochemistry (Mosc). 2012 Feb;77(2):165-70. doi: 10.1134/S0006297912020071.
We earlier described CBS-pyrophosphatase of Moorella thermoacetica (mtCBS-PPase) as a novel phosphohydrolase that acquired a pair of nucleotide-binding CBS domains during evolution, thus endowing the protein with the capacity to be allosterically regulated by adenine nucleotides (Jämsen, J., Tuominen, H., Salminen, A., Belogurov, G. A., Magretova, N. N., Baykov, A. A., and Lahti, R. (2007) Biochem. J., 408, 327-333). We herein describe a more evolved type of CBS-pyrophosphatase from Clostridium perfringens (cpCBS-PPase) that additionally contains a DRTGG domain between the two CBS domains in the regulatory part. cpCBS-PPase retained the ability of mtCBS-PPase to be inhibited by micromolar concentrations of AMP and ADP and activated by ATP and was additionally activated by diadenosine polyphosphates (AP(n)A) with n > 2. Stopped-flow measurements using a fluorescent nucleotide analog, 2'(3')-O-(N-methylanthranoyl)-AMP, revealed that cpCBS-PPase interconverts through two different conformations with transit times on the millisecond scale upon nucleotide binding. The results suggest that the presence of the DRTGG domain affords greater flexibility to the regulatory part, allowing it to more rapidly undergo conformational changes in response to binding.
我们之前曾描述过 Moorella thermoacetica（mtCBS-PPase）的 CBS-焦磷酸酶是一种新型的磷酸水解酶，在进化过程中获得了一对核苷酸结合 CBS 结构域，从而使该蛋白具有被腺嘌呤核苷酸别构调节的能力（Jämsen，J.，Tuominen，H.，Salminen，A.，Belogurov，G. A.，Magretova，N. N.，Baykov，A. A.，和 Lahti，R.（2007）生物化学杂志，408，327-333）。我们在此描述了来自梭状芽孢杆菌（cpCBS-PPase）的一种进化程度更高的 CBS-焦磷酸酶，其在调节部分的两个 CBS 结构域之间还含有 DRTGG 结构域。cpCBS-PPase保留了 mtCBS-PPase 被微摩尔浓度的 AMP 和 ADP 抑制以及被 ATP 激活的能力，并且还被二腺苷多磷酸（AP(n)A）激活，其中 n > 2。使用荧光核苷酸类似物 2'（3'）-O-（N-甲基蒽酰）-AMP 的停流测量显示，cpCBS-PPase在核苷酸结合时通过两种不同的构象进行互变，转换时间在毫秒范围内。结果表明，DRTGG 结构域的存在使调节部分具有更大的灵活性，使其能够更快地响应结合而发生构象变化。
