Selective dissociation of non-covalent bonds in biological molecules by laser spray.
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.
Takamizawa
Atsushi
A
Clean Energy Research Center, University of Yamanashi, Takeda-4, Kofu 400-8511, Japan.
Itoh
Yoshiyuki
Y
Osawa
Ryo
R
Iwasaki
Noriyuki
N
Nishimura
Yoshifumi
Y
Akashi
Satoko
S
Hiraoka
Kenzo
K
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D004247:DNA / Q000737:chemistry* / Q000378:metabolism* / Q000528:radiation effects; D007259:Infrared Rays; D007834:Lasers; D008969:Molecular Sequence Data; D011485:Protein Binding / Q000528:radiation effects; D011487:Protein Conformation / Q000528:radiation effects; D011489:Protein Denaturation / Q000528:radiation effects; D017510:Protein Folding; D020598:Proto-Oncogene Proteins c-myb / Q000737:chemistry* / Q000378:metabolism* / Q000528:radiation effects; D012710:Serum Albumin, Bovine / Q000737:chemistry* / Q000528:radiation effects; D012996:Solutions / Q000737:chemistry / Q000528:radiation effects; D021241:Spectrometry, Mass, Electrospray Ionization
D020598:Proto-Oncogene Proteins c-myb; D012996:Solutions; D012710:Serum Albumin, Bovine; D004247:DNA