Metal-chelating plastic MALDI (pMALDI) chips for the enhancement of phosphorylated-peptide/protein signals.
A disposable polymeric pMALDI array with a universal metal cation-chelatable surface for pretreatment/signal enhancement of phosphoproteins and/or phosphopeptides in complex samples was developed. Acrylic acid N-hydroxysuccinimide ester and methyl methacrylate monomers were copolymerized in thin layer molds in a 1:13.3 molar ratio and subsequently treated with Nalpha,Nalpha-bis(carboxymethyl)-l-lysine to obtain a structured planar MALDI array. The prepared NTA pMALDI chip array was activated with metal cations (e.g., Ga(III), Ni(II)), and the selectivities for phosphopeptides (e.g., trypsin-digested alpha-casein (alpha-Cas), and phospho-angiotensin II (p-Ang)) were evaluated using MALDI-TOF/MS. The highest selectivity for proteins was observed for the Ni(II)-NTA chip. The p-Ang was enriched in the presence of BSA tryptic peptides ca. 5 times and represented the major peak after sample adsorption/washing on Ga(III)-NTA chip. The performance of the Ga(III)-chip, tested on alpha-Cas tryptic digest, is fully comparable to commercial systems. Additionally, higher MW peptides and limited methionine oxidation were observed with the chip. A combination of selective absorption of phosphoproteins on Ni(II)-chips and the further enrichment of digested phosphopeptides on the Ga(III)-chip can prove to be very useful for fast identification of unknown proteins using MALDI-TOF/MS.
Ibañez
Alfredo J
AJ
Mass Spectrometry Research Group, Max Planck Institute for Chemical Ecology, Hans-Knöll-Strasse 8, 07745 Jena, Germany.
Muck
Alexander
A
Svatos
Ales
A
D000327:Adsorption; D000804:Angiotensin II / Q000737:chemistry; D005708:Gallium / Q000737:chemistry; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D010449:Peptide Mapping; D010455:Peptides / Q000737:chemistry*; D010748:Phosphopeptides; D010750:Phosphoproteins / Q000737:chemistry; D010766:Phosphorylation; D011108:Polymers / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D040901:Proteomics / Q000295:instrumentation* / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D010455:Peptides; D010748:Phosphopeptides; D010750:Phosphoproteins; D011108:Polymers; D011506:Proteins; D000804:Angiotensin II; D005708:Gallium