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Quantification of Protein-Ligand Interactions by Laser Electrospray Mass Spectrometry
Journal of the American Society for Mass Spectrometry ( IF 3.1 ) Pub Date : 2018-04-13 , DOI: 10.1007/s13361-018-1935-9
Jieutonne J. Archer 1 , Santosh Karki 1 , Fengjian Shi 1 , Habiballah Sistani 1 , Robert J. Levis 1
Affiliation  

Laser electrospray mass spectrometry (LEMS) measurement of the dissociation constant (Kd) for hen egg white lysozyme (HEWL) and N,N′,N″-triacetylchitotriose (NAG3) revealed an apparent Kd value of 313.2 ± 25.9 μM for the ligand titration method. Similar measurements for N,N′,N″,N″’-tetraacetylchitotetraose (NAG4) revealed an apparent Kd of 249.3 ± 13.6 μM. An electrospray ionization mass spectrometry (ESI-MS) experiment determined a Kd value of 9.8 ± 0.6 μM. In a second LEMS approach, a calibrated measurement was used to determine a Kd value of 6.8 ± 1.5 μM for NAG3. The capture efficiency of LEMS was measured to be 3.6 ± 1.8% and is defined as the fraction of LEMS sample detected after merging with the ESI plume. When the dilution is factored into the ligand titration measurement, the adjusted Kd value was 11.3 μM for NAG3 and 9.0 μM for NAG4. The calibration method for measuring Kd developed in this study can be applied to solutions containing unknown analyte concentrations.

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Graphical Abstract



中文翻译:

蛋白质-配体相互作用的激光电喷雾质谱定量

解离常数(K的激光电喷雾质谱法(LEMS)测量d为鸡蛋白溶菌酶(HEWL)和)ÑÑ ',Ñ “-triacetylchitotriose(NAG 3)揭示的表观ķ d 313.2±25.9μM的值对配体滴定法。类似测量为ÑÑ ',Ñ “,Ñ ”'-tetraacetylchitotetraose(NAG 4)揭示的表观ķ d 249.3±13.6微米。电喷雾电离质谱(ESI-MS)实验确定了K d值为9.8±0.6μM。在第二种LEMS方法中,使用校准的测量确定NAG 3的K d值为6.8±1.5μM 。测得LEMS的捕获效率为3.6±1.8%,定义为与ESI羽流合并后检测到的LEMS样品的分数。当将稀释液计入配体滴定测量时,对于NAG 3而言,调整后的K d值为11.3μM,对于NAG 4而言,调整后的K d值为9.0μM 。本研究中开发的用于测量K d的校准方法可以应用于含有未知分析物浓度的溶液。

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更新日期:2018-04-13
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