Ion soft-landing into liquids: Protein identification, separation, and purification with retention of biological activity.
Protein ions, after mass spectrometric separation, can be soft-landed into liquid surfaces with preservation of their native structures. Retention of biological activity is strongly favored in glycerol-based surfaces but not in self-assembled monolayer solid surfaces. Soft-landing efficiency for multiply-charged hexokinase ions was found to be some four times higher for a glycerol/fructose liquid surface than for a fluorinated self-assembled monolayer surface. Soft-landing into liquid surfaces is also shown to allow (1) protein purification, (2) on-surface identification of the soft-landed material using MALDI, and (3) protein identification by in-surface tryptic digestion. Pure lysozyme was successfully isolated from different mixtures including an oxidized, partially decomposed batch of the protein and a partial tryptic digest. Liquid glycerol/carbohydrate mixtures could be used directly to record MALDI spectra on the soft-landed compounds provided they were fortified in advance with traditional MALDI matrices such as p-nitroaniline and alpha-cyano-4-hydroxycinnamic acid. Various proteins were soft-landed and detected on-target using these types of liquid surface. Soft-landing of multiply-charged lysozyme ions onto fluorinated self-assembled monolayer surfaces was found to occur with a limited amount of neutralization, and trapped multiply-charged ions could be desorbed from the surface by laser desorption. Initial data is shown for a new approach to protein identification that combines top-down and bottom-up approaches by utilizing protein ion soft-landing from a protein mixture, followed by tryptic digestion of the landed material and detection of characteristic tryptic fragments by MALDI.
Gologan
Bogdan
B
Department of Chemistry, Purdue University, West Lafayette, IN 47907-1393, USA.
Takáts
Zoltán
Z
Alvarez
Jormarie
J
Wiseman
Justin M
JM
Talaty
Nari
N
Ouyang
Zheng
Z
Cooks
R Graham
RG
D000595:Amino Acid Sequence; D006593:Hexokinase / Q000032:analysis; D007477:Ions; D008969:Molecular Sequence Data; D009113:Muramidase / Q000032:analysis; D010455:Peptides / Q000032:analysis*; D011506:Proteins / Q000032:analysis*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000295:instrumentation / Q000379:methods*
D007477:Ions; D010455:Peptides; D011506:Proteins; D006593:Hexokinase; D009113:Muramidase