Rhyu G I, Ray W J, Markley J L
Biochemistry. 1985 May 7;24(10):2536-41. doi: 10.1021/bi00331a021.
Metal binding at the activating site of rabbit muscle phosphoglucomutase has been studied by 31P, 7Li, and 113Cd NMR spectroscopy. A 7Li NMR signal of the binary Li+ complex of the phosphoenzyme was not observed probably because of rapid transverse relaxation of the bound ion due to chemical exchange with free Li+. The phosphoenzyme-Li+-glucose 6-phosphate ternary complex is more stable, kinetically, and yields a well-resolved peak from bound Li+ at -0.24 ppm from LiCl with a line width of 5 Hz and a T1 relaxation time of 0.51 +/- 0.07 s at 78 MHz. When glucose 1-phosphate was bound, instead, the chemical shift of bound 7Li+ was -0.13 ppm; and in the Li+ complex of the dephosphoenzyme and glucose bisphosphate a partially broadened 7Li+ peak appeared at -0.08 ppm. Thus, the bound metal ion has a somewhat different environment in each of these three ternary complexes. The 113Cd NMR signal of the binary Cd2+ complex of the phosphoenzyme appears at 22 ppm relative to Cd(ClO4)2 with a line width of 20 Hz at 44.4 MHz. Binding of substrate and formation of the Cd2+ complex of the dephosphoenzyme and glucose bisphosphate broaden the 113Cd NMR signal to 70 Hz and shift it to 75 ppm. The 53 ppm downfield shift upon the addition of substrate along with 1H NMR data suggests that one oxygen ligand to Cd2+ in the binary complex is replaced by a nitrogen ligand at some intermediate point in the enzymic reaction.(ABSTRACT TRUNCATED AT 250 WORDS)
通过³¹P、⁷Li和¹¹³Cd核磁共振光谱研究了兔肌肉磷酸葡萄糖变位酶活性位点处的金属结合情况。未观察到磷酸化酶的二元Li⁺复合物的⁷Li核磁共振信号，这可能是由于结合离子与游离Li⁺发生化学交换导致横向弛豫迅速。从动力学角度看，磷酸化酶-Li⁺-6-磷酸葡萄糖三元复合物更稳定，在78兆赫兹时，结合的Li⁺产生了一个分辨率良好的峰，相对于LiCl，其化学位移为-0.24 ppm，线宽为5赫兹，T1弛豫时间为0.51±0.07秒。相反，当结合1-磷酸葡萄糖时，结合的⁷Li⁺的化学位移为-0.13 ppm；在脱磷酸化酶和双磷酸葡萄糖的Li⁺复合物中，一个部分变宽的⁷Li⁺峰出现在-0.08 ppm处。因此，在这三种三元复合物中，结合的金属离子所处环境略有不同。磷酸化酶的二元Cd²⁺复合物的¹¹³Cd核磁共振信号相对于Cd(ClO₄)₂出现在22 ppm处，在44.4兆赫兹时线宽为20赫兹。底物的结合以及脱磷酸化酶和双磷酸葡萄糖的Cd²⁺复合物的形成使¹¹³Cd核磁共振信号变宽至70赫兹，并将其位移至75 ppm。添加底物时产生的53 ppm的向下位移以及¹H核磁共振数据表明，在酶促反应的某个中间点，二元复合物中与Cd²⁺结合的一个氧配体被一个氮配体取代。（摘要截选至250字）
