Accelerated proteolysis in alternating electric fields for peptide mapping.
Sinusoidal alternating voltages (typically 5 V) were employed to enhance the efficiency of proteolysis for peptide mapping in this work. Protein solutions containing trypsin were allowed to digest with the assistance of alternating electric fields (AEFs) between a pair of platinum wire electrodes in Eppendorf tubes. The feasibility and performance of the novel proteolysis approach were investigated by the digestion of several standard proteins. It was demonstrated that AEFs significantly accelerated in-solution proteolysis and the digestion time was substantially reduced to 5 min. The digests were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) with sequence coverages that were comparable to those obtained by using conventional 12-h in-solution proteolysis. The suitability of AEF-assisted proteolysis to real protein samples was demonstrated by digesting and identifying human serum albumin in gel separated from human serum by sodium dodecyl sulphate/polyacrylamide gel electrophoresis (SDS-PAGE). The present proteolysis strategy is simple and efficient and will find a wide range of applications in protein identification.
Wang
Sheng
S
School of Pharmacy & Department of Chemistry, Fudan University, Shanghai 200032, China.
Bao
Huimin
H
Liu
Ting
T
Zhang
Luyan
L
Yang
Pengyuan
P
Chen
Gang
G
D004574:Electromagnetic Fields; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007202:Indicators and Reagents; D010449:Peptide Mapping / Q000379:methods*; D010455:Peptides / Q000737:chemistry*; D011492:Protein Hydrolysates / Q000032:analysis; D012709:Serum Albumin / Q000737:chemistry; D012996:Solutions; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014357:Trypsin / Q000737:chemistry
D007202:Indicators and Reagents; D010455:Peptides; D011492:Protein Hydrolysates; D012709:Serum Albumin; D012996:Solutions; D014357:Trypsin