McKillop, Aine, Meade, A, Flatt, Peter and O'Harte, Finbarr (2003) Evaluation of the site(s) of glycation in human proinsulin by ion-trap LCQ electrospray ionization mass spectrometry. REGULATORY PEPTIDES, 113 (1-3). pp. 1-8. [Journal article]
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The glycation of beta cell proteins is known to occur under hyperglycemic states. The site(s) of glycation in human proinsulin was investigated following exposure to a hyperglycemic environment under reducing conditions in vitro. Proinsulin and glycated proinsulin were separated by reversed-phase high-performance liquid chromatography (R-P-HPLC) and identified using LCQ ion-trap electrospray ionization mass spectrometry. This revealed a major peak (>70% total) of monoglycated proinsulin (M-r 9552.2 Da), a second peak (approximately 27%) of nonglycated proinsulin (M-r 9389.8 Da), and a third minor peptide peak (approximately 3%) corresponding to diglycated proinsulin (M-r 9717.9 Da). Following reduction of disulphide bridges with dithiothreitol, intact peptides were incubated with endoproteinase Glu-C to release nine daughter fragments for LC-MS analysis. This strategy revealed an N-terminal fragment of monoglycated proinsulin Phe(1) Glu(13), which contained a single glucitol adduct (M-r 1642.0 Da). A similar treatment of small amounts of purified diglycated proinsulin revealed a fragment with Phe(1)-Glu(13) linked by a disulphide bridge to Gln(70)-Glu(82) containing two glucitol adducts (M-r 3292.7 Da). In summary, these studies indicate that the major site of glycation in proinsulin, like insulin, is the amino terminal Phe(1) residue. However, small amounts of diglycated proinsulin occur naturally, involving an additional site of glycation located between Gln(70) and Glu(82). (C) 2002 Elsevier Science B.V. All rights reserved.
|Item Type:||Journal article|
|Faculties and Schools:||Faculty of Life and Health Sciences|
Faculty of Life and Health Sciences > School of Biomedical Sciences
|Research Institutes and Groups:||Biomedical Sciences Research Institute|
Biomedical Sciences Research Institute > Diabetes
|Deposited By:||Professor Peter Flatt|
|Deposited On:||14 Jan 2010 15:56|
|Last Modified:||28 Feb 2011 11:16|
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