Science and Technology Production
Book Chapter
Authorship
S. L. Giudicessi
;
M. L. Salum
;
M. C. Martínez-Ceron
;
O. Cascone
;
R. Erra-Balsells
;
CAMPERI, SILVIA ANDREA
Date
2015
Publishing House and Editing Place
American Peptide Society
Book
Peptides 2015 Proceedings of the Twenty-Fourth American Peptide Symposium
(pp. 131-133)
American Peptide Society
American Peptide Society
ISBN
978-0-9839741-5-4
Summary
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SIGEVA
Short peptides are widely used as ligands in affinity chromatography purification of proteins. However, proteases present in the crude sample may degrade immobilized peptides, shortening the chromatographic matrix useful life. Then, peptide ligand stability must be evaluated before its use in a purification process. Commonly, enzymatic stability is evaluated with soluble peptides, which differs from the immobilized peptide behavior. Further, the study of the peptide degradation products require...
Short peptides are widely used as ligands in affinity chromatography purification of proteins. However, proteases present in the crude sample may degrade immobilized peptides, shortening the chromatographic matrix useful life. Then, peptide ligand stability must be evaluated before its use in a purification process. Commonly, enzymatic stability is evaluated with soluble peptides, which differs from the immobilized peptide behavior. Further, the study of the peptide degradation products requires purification steps before analysis. In this work we developed a method to evaluate immobilized peptide stability using MALDI-MS or ESI-MS. ChemMatrix was used as immobilized support due to its chemical stability. This PEG-based matrix allowed peptide synthesis in organic solvents and stability peptide evaluation in aqueous solvents. 4-hydroxymethylbenzoic acid was used as linker. The model peptide FKFRYTAHSGASG was synthesized using the Fmoc strategy. Peptide-beads were then incubated with solution containing proteases. After washing, the beads peptides were detached with ammonia vapor and analyzed by MS and MS/MS. Both ESI-MS and MALDI-MS allowed the detection of the whole peptide as well as their C-terminal enzymatic degradation products. Although E--cyano-4-hydroxycinnamic acid matrix clusters interfered in MALDI-MS analysis of low molecular weight products, Z-sinapinic acid matrix allowed their analysis.
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Key Words
MS/MSstabilityPeptideEnzimes