Denaturation of alpha-lactalbumin and ubiquitin studied by electrospray and laser spray.
Electrospray and laser spray mass spectra of human alpha-lactalbumin and bovine ubiquitin were studied, with an emphasis on the denaturation induced by laser spray. There were no remarkable differences in the electrospray and laser spray mass spectra for acidic and basic aqueous solutions of alpha-lactalbumin in positive and negative modes of operations. This originates from the fact that this protein is tightly folded with four disulfide bonds. For ubiquitin, however, denaturation was induced by laser spray for the positive mode of operation and the [M+nH](n+) with a maximum of n = 13 was observed, i.e., all the acidic amino acid residues are fully neutralized (protonated). In contrast, the laser-induced denaturation was not observed for the negative mode of operation, i.e., denaturation of ubiquitin is largely suppressed in the negatively charged liquid droplets. The marked difference observed in the positive and negative modes of operations for ubiquitin is ascribed to the difference in the susceptibility of side-chain/main-chain interactions in the positive-ion excess and in the negative-ion excess liquid droplets. That is, the interactions between the basic residues and main-chain amide carbonyl groups (-NH(3) (+)***O=C< or -NH(2)***O=C<) which play an important role in stabilizing the protein structures are not so affected in the negative mode of operation but are weakened in the positive mode of operation.
Nakamura
Masana
M
Clean Energy Research Center, University of Yamanashi, Takeda-4, Kofu 400-8511, Japan.
Takamizawa
Atsushi
A
Yamada
Hideki
H
Hiraoka
Kenzo
K
Akashi
Satoko
S
D030562:Databases, Protein; D006801:Humans; D007768:Lactalbumin / Q000737:chemistry*; D011487:Protein Conformation; D011489:Protein Denaturation; D012997:Solvents; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D025801:Ubiquitin / Q000737:chemistry*
D012997:Solvents; D025801:Ubiquitin; D007768:Lactalbumin