Selective dissociation of non-covalent bonds in biological molecules by laser spray.
Selective dissociation of non-covalent bonds in biological molecules by laser spray.
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DOI：
10.1002/jms.677
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发表时间：
2004-09
期刊：
影响因子：
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通讯作者：
Atsushi Takamizawa;Yoshiyuki Itoh;R. Osawa;Noriyuki Iwasaki;Y. Nishimura;S. Akashi;K. Hiraoka
中科院分区：
文献类型：
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作者：
Atsushi Takamizawa;Yoshiyuki Itoh;R. Osawa;Noriyuki Iwasaki;Y. Nishimura;S. Akashi;K. Hiraoka
本实验室研制的激光喷雾法用于牛血清白蛋白（BSA）、双链DNA（dsDNA）和蛋白质-DNA复合物的分析。通过将激光功率从0 W（电喷雾）增加到1.4W，用10.6 μ m红外激光照射不锈钢毛细管的尖端。激光束在不锈钢毛细管的尖端处聚焦至约0.3mm。当BSA水溶液被激光照射时，除了非变性单体、二聚体和三聚体部分的多电荷离子外，还新观察到高电荷单体离子。这表明BSA在溶液中受到红外激光照射时发生变性。在本实验条件下，1.4W的激光功率不足以使BSA完全变性。而双链DNA被发现几乎完全解离的单链DNA成分的激光照射功率为1.2 W，没有观察到DNA分子的片段化。对于蛋白质-DNA复合物，即c-Myb DNA结合结构域和dsDNA的复合物，观察到复合物解离成组分部分。这些发现表明激光喷雾可以选择性地将非共价复合物解离成亚基，而不引起亚基的共价键的解离。激光喷雾将成为研究生物分子（包括非共价复合物）结构和稳定性的一种通用方法。
The laser spray developed in our laboratory was applied to the analysis of bovine serum albumin (BSA), double-stranded DNA (dsDNA) and a protein-DNA complex. The tip of a stainless-steel capillary was irradiated with a 10.6 micro m infrared laser by increasing the laser power from 0 W (electrospray) to 1.4 W. The laser beam was focused to about 0.3 mm at the tip of the stainless-steel capillary. When BSA aqueous solution was irradiated by the laser, highly charged monomer ions were newly observed in addition to the multiply charged ions of non-denatured monomer, dimer and trimer moieties. This indicates that BSA suffers from denaturation on irradiation with an infrared laser in solution. A 1.4 W laser power is not sufficient to cause the complete denaturation of BSA under the present experimental conditions. Whereas dsDNA was found to dissociate almost completely to single-stranded DNA constituents on laser irradiation with a power of 1.2 W, no fragmentation of DNA molecules was observed. For a protein-DNA complex, i.e. a complex of c-Myb DNA binding domain and dsDNA, dissociation of the complex to the component moieties was observed. These findings indicate that the laser spray can selectively dissociate non-covalent complexes into subunits without causing dissociation of the covalent bonds of the subunits. The laser spray will be a versatile method for the investigation of the structures and stabilities of biomolecules including non-covalent complexes.