Electrochemical analysis of Os(VI)-modified glycoproteins and label-free glycoprotein detection eluted from lectin capillary column. (10th June 2017)

• Record Type: Journal Article
• Authors: Trefulka, Mojmír; Dorčák, Vlastimil; Křenková, Jana; Foret, František; Paleček, Emil
• Is Part Of: Electrochimica acta. Volume 239(2017)
• Page Start: 10
• Page End: 15
• Publication Date: 2017-06-10
• Subjects: Glycan chemical modification in RNase B -- Electrochemical glycoprotein detection -- Mercury and carbon electrodes -- Voltammetry and constant-current chronopotentiometry -- Lectin monolithic flow-through column; Electrochemistry -- Periodicals; Electrochemistry, Industrial -- Periodicals; 541.37
• DOI: 10.1016/j.electacta.2017.04.045
• Languages: English
• ISSNs: 0013-4686

Abstract

Graphical abstract: Highlights: Glycoprotein modification by Os(VI) complexes. Determination of Os(VI)-modified RNase B at pM concentrations. Differences in stabilities of Os(VI)-modified and unmodified RNases at electrodes. Label-free chronopotentiometric glycoprotein detection eluted from a lectin column. Abstract: Earlier we showed that using Os(VI)temed complex, glycans can be modified directly in glycoproteins and detected voltammetrically at μM concentrations at carbon electrodes. Here we used Os(VI)2, 2′-bipyridine for modification of ribonucleases and Hg electrodes for voltammetric detection. We show that glycosylated RNase B produced electrocatalytic signal (after separation from the reaction mixture) at pM concentrations while non-glycosylated RNase A yielded negligible signal under the same conditions. Using Os(VI)temed and carbon electrodes voltammetric detection was less sensitive but allowed detection of RNase B in an excess of non-glycosylated protein, directly in the reaction mixture. We also showed that the constant current chronopotentiometric stripping (CPS) peak H (which is due to the ability of some amino acid residues in proteins to catalyze hydrogen evolution at Hg electrodes) could be used for protein structure-sensitive analysis at mercury electrodes. Using this peak, here we show that glycosylation of RNase stabilizes its molecule at the electrode. On the other hand, Os(VI)temed modification results in destabilization of this surface-attached protein. Peak H was also used for detection of RNase B separated from a large excess of non-glycosylated proteins on a lectin (concanavalin A) monolithic flow-through column.