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Yang, J.Z.* ; Röwer, C.* ; Koy, C.* ; Ruß, M.* ; Rüger, C.P. ; Zimmermann, R. ; Fritschen, U.V.* ; Bredell, M.* ; Finke, J.C.* ; Glocker, M.O.*

Mass spectrometric characterization of limited proteolysis activity in human plasma samples under mild acidic conditions.

Methods 89, 30-37 (2015)
Verlagsversion DOI
Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.
We developed a limited proteolysis assay for estimating dynamics in plasma-borne protease activities using MALDI ToF MS analysis as readout. A highly specific limited proteolysis activity was elicited in human plasma by shifting the pH to 6. Mass spectrometry showed that two singly charged ion signals at m/z 2753.44 and m/z 2937.56 significantly increased in abundance under mild acidic conditions as a function of incubation time. For proving that a provoked proteolytic activity in mild acidic solution caused the appearance of the observed peptides, control measurements were performed (i) with pepstatin as protease inhibitor, (ii) with heat-denatured samples, (iii) at pH 1.7, and (iv) at pH 7.5. Mass spectrometric fragmentation analysis showed that the observed peptides encompass the amino acid sequences 1-24 and 1-26 from the N-terminus of human serum albumin. Investigations on peptidase specificities suggest that the two best candidates for the observed serum albumin cleavages are cathepsin D and E. Reproducibility, robustness, and sensitivity prove the potential of the developed limited proteolysis assay to become of clinical importance for estimating dynamics of plasma-borne proteases with respect to associated pathophysiological tissue conditions.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Schlagwörter Limited Proteolysis ; Maldi Tof Ms ; Plasma Proteins ; Protein Structure ; Tissue Quality Assay
ISSN (print) / ISBN 1046-2023
e-ISSN 1095-9130
Zeitschrift Methods
Quellenangaben Band: 89, Heft: , Seiten: 30-37 Artikelnummer: , Supplement: ,
Verlag Elsevier
Verlagsort Amsterdam [u.a.]
Begutachtungsstatus Peer reviewed