APPLICATION OF ELECTROSPRAY MASS-SPECTROMETRY IN PROBING PROTEIN-PROTEIN AND PROTEIN LIGAND NONCOVALENT INTERACTIONS
APPLICATION OF ELECTROSPRAY MASS-SPECTROMETRY IN PROBING PROTEIN-PROTEIN AND PROTEIN LIGAND NONCOVALENT INTERACTIONS
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DOI：
10.1016/1044-0305(93)85026-t
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发表时间：
1993-08-01
影响因子：
3.2
通讯作者：
COVEY, TR
中科院分区：
文献类型：
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作者：
HUANG, EC;PRAMANIK, BN;COVEY, TR
描述了一种基于电喷雾电离(ESI)的新的质谱学方法，用于检测蛋白质-蛋白质[干扰素-伽马二聚体]和蛋白质-配体[ras-鸟苷二磷酸(GDP)]的非共价相互作用。该方法利用水溶液中的电喷雾，在适当的pH下进行测定。观察到的平均相对分子质量为33,819 Da，证实了干扰素-γ二聚体非共价复合体的存在。检测干扰素-伽马二聚体的关键是使用碱性溶液(pH约为9)进行样品制备和质谱分析。还研究了离子采样孔和聚焦四极杆区域的离团能量对气相非共价复合体(干扰素-伽马二聚体)保存的影响。进一步扩展了离聚能对络合物解离的影响，探讨了ras-GDP的非共价蛋白质-配体缔合作用。研究发现，解离干扰素-伽马二聚体需要很少的能量，而解离气相ras-GDP复合体需要相当大的能量。
A novel mass spectrometry-based methodology using electrospray ionization (ESI) is described for the detection of protein-protein [interferon (IFN)-gamma dimer] and protein-ligand [ ras-guanosine diphosphate (GDP)] noncovalent interactions. The method utilizes ESI from aqueous solution at appropriate pH. The presence of the noncovalent complex of the IFN-gamma dimer was confirmed by the observed average molecular weight of 33,819 Da. The key to the detection of the IFN-gamma dimer is the use of an alkaline solution (pH approximately 9) for sample preparation and for mass spectrometry analysis. The effect of the declustering energy in the region of the ion sampling orifice and focusing quadrupole on the preservation of the gas-phase noncovalent complex (IFN-gamma dimer) was also studied. The effect of the declustering energy on complex dissociation was further extended to probe the noncovalent protein-ligand association of ras-GDP. It was found that little energy is required to dissociate the IFN-gamma dimer, whereas a substantial amount of energy is required to dissociate the gas-phase ras-GDP complex.