The calcium-modulated structures of calmodulin and S100b proteins are useful to monitor hydrogen/deuterium exchange efficiency using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.
Hydrogen/deuterium exchange (HDX) using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF) is a sensitive, salt-tolerant and high-throughput method useful to probe protein conformation and molecular interactions. However, a drawback of the MALDI HDX technique is that sample preparation methods can typically result in higher levels of artificial deuterium in-exchange and/or hydrogen back- exchange just prior to or during mass analysis; this may impair data reproducibility and impede structural and kinetic data interpretation. While methods to minimize effects of back-exchange during protein analyte deposition on MALDI plates have been reported, this study presents a readily available, highly sensitive protein control set to facilitate rapid MALDI HDX protocol workup. The Ca(2+)-induced solvent accessible surface area (ASA) changes of calmodulin (CaM) and S100 proteins were employed to monitor and optimize HDX protocol efficiency. Under non- stringent room temperature conditions, the Ca(2+)-induced deuterium exchange of CaM, DeltaD(ca2+ -apo), MH(+) shifts -17 to -24 Da, while S100 DeltaD(ca2+ -apo) MH(+) shifts +8 to +12 Da. By comparing the divergent CaM and S100 Ca(2+)-induced deuterium mass shift differences, HDX sample workup and MALDI plate spotting conditions can easily be monitored.
Pingerelli
Peter L
PL
Western International University, 9215 N Black Canyon Hwy, Phoenix, AZ 85021, USA. ppingerelli@mywiu.wintu.edu
Ozols
Victor V
VV
Saleem
Haroon
H
Anderson
Carly R
CR
Burns
Richard S
RS
D002147:Calmodulin / Q000737:chemistry*; D041961:Deuterium Exchange Measurement / Q000379:methods*; D006801:Humans; D009414:Nerve Growth Factors / Q000737:chemistry*; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D002147:Calmodulin; D009414:Nerve Growth Factors; D011994:Recombinant Proteins; D064568:S100 Calcium Binding Protein beta Subunit; D009418:S100 Proteins; C579160:S100B protein, human