Substrate specificity for catalysis of phosphoryl transfer by the calcium ATPase of sarcoplasmic reticulum.
Substrate specificity for catalysis of phosphoryl transfer by the calcium ATPase of sarcoplasmic reticulum.
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肌浆网钙 ATP 酶催化磷酰基转移的底物特异性。
DOI：
10.1006/abbi.1994.1355
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发表时间：
1994
影响因子：
3.9
通讯作者：
Jencks,WP
中科院分区：
文献类型：
--
作者：
Myung,J;Jencks,WP
当将α，β-亚甲基ADP（α，β-CH 2-ADP）加入到与Ca 2+结合的肌浆网磷酸化钙ATP酶中时，不合成Ca 2· E β P · Mg，α，β-亚甲基ATP（5 mM MgCl 2，100 mM KCl，pH 7.0，25°C）。类似地，腺苷5′-O-（2-硫代三磷酸）不是由磷酸酶与腺苷5′-O-（2-硫代二磷酸）（ADPβS）反应合成的。相反，ATP是由磷酸酶与ADP反应快速可逆地形成的。两种ADP类似物对ADP与磷酸酶的结合都是竞争性抑制剂，KADPS = 0.45mM：α，β-CH_2-ADP和ADPβS与磷酸酶的结合分别为K α，β-CH_2-ADPS= 0.92mM和KADP βSS= 0.05mM。我们得出的结论是，从磷酸酶到α，β-CH 2-ADP的磷酰基转移在动力学上被阻断，尽管它在热力学上是有利的。与α，β-CH 2-ADP相比，Ca 2· E·P · Mg向ADP的磷酰基转移速率加快> 105，这可能是由于ADP与α，β-CH 2-ADP在结构和净电荷上的差异所致。从磷酸酶到ADPβS的磷酰基转移在化学上是如此不利，以至于我们不能确定过渡态是否也是不利的。
When α,β-methylene ADP (α,β-CH2-ADP) is added to the phosphorylated calcium ATPase of sarcoplasmic reticulum with Ca2+-bound, Ca2· E ∼ P · Mg, α,β-methylene ATP is not synthesized (5 mM MgCl2, 100 mM KCl, pH 7.0, 25°C). Similarly, adenosine 5′-O-(2-thiotriphosphate) is not synthesized from reaction of the phosphoenzyme with adenosine 5′-O-(2-thiodiphosphate), ADPβS. In contrast, ATP is formed rapidly and reversibly from the reaction of the phosphoenzyme with ADP. Both ADP analogs are competitive inhibitors for the binding of ADP to the phosphoenzyme withKADPS= 0.45 mM: α,β-CH2-ADP and ADPβS bind to the phosphoenzyme withKα,β-CH2-ADPS= 0.92 mM andKADPβSS= 0.05 mM, respectively. We conclude that phosphoryl transfer from the phosphoenzyme to α,β-CH2-ADP is kinetically blocked, although it is thermodynamically favorable. The rate acceleration of >105for phosphoryl transfer from Ca2· E ∼ P · Mg to ADP compared to α,β-CH2-ADP can be attributed to the differences in both the structure and the net charge of ADP compared with α,β-CH2-ADP at pH 7.0. Phosphoryl transfer from the phosphoenzyme to ADPβS is thermodynamically so unfavorable that we cannot determine whether the transition state is also unfavorable.