Protein identification with Teflon as matrix-assisted laser desorption/ionization sample support.
Protein identification is a critical step in proteomics, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) plays an important role in that identification. Polytetrafluoroethylene (Teflon) was tested as a new MALDI sample support to improve protein identification. The tryptic peptides obtained from a model protein were bound to the surface of a modified MALDI sample holder via the hydrophobic interactions that occur between the Teflon surface and the peptide ion-pairs, and the affinity of alpha-cyano-4-hydroxycinnamic acid for the peptides. During that surface-binding step, the peptide mixture was also desalted and concentrated. A greater number of matched peptides and a larger sequence coverage were obtained for the proteins when Teflon was used as the sample support compared with conventional sample preparation methods and a stainless-steel surface. In addition, the characterization of a small amount of protein was improved with Teflon. Nine silver-stained protein spots obtained from 2-D gel of a human cerebrospinal fluid (CSF) proteome were identified by this method. Among the nine protein spots, peptide 6:c3c fragment and procollagen c-proteinase enhancer were not annotated in any published 2-D map of human CSF. A Teflon MALDI sample support is a low-cost, simple, and effective method that can be used to improve the quality of the MALDI mass spectrum of a complex tryptic peptide mixture, and to achieve a higher level of reliability and success in protein identification.
Yuan
Xianglin
X
Charles B. Stout Neuroscience Mass Spectrometry Laboratory University of Tennessee Health Science Center, 847 Monroe Avenue, Memphis, Tennessee 38163, USA.
Desiderio
Dominic M
DM
D000818:Animals; D002417:Cattle; D002556:Cerebrospinal Fluid Proteins / Q000032:analysis*; D015180:Electrophoresis, Gel, Two-Dimensional; D004867:Equipment Design; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D017116:Low Back Pain / Q000134:cerebrospinal fluid; D010449:Peptide Mapping; D011138:Polytetrafluoroethylene; D020543:Proteome; D012680:Sensitivity and Specificity; D012710:Serum Albumin, Bovine / Q000737:chemistry; D016622:Silver Staining; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000295:instrumentation* / Q000379:methods; D013193:Stainless Steel; D013499:Surface Properties; D014357:Trypsin / Q000494:pharmacology
D002556:Cerebrospinal Fluid Proteins; D020543:Proteome; D013193:Stainless Steel; D012710:Serum Albumin, Bovine; D011138:Polytetrafluoroethylene; D014357:Trypsin