α1-Microglobulin chromophores are located to three lysine residues semiburied in the lipocalin pocket and associated with a novel lipophilic compound
| Type: | Application |
Scientific Paper |
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| Number: | Technology |
L01:1:149 |
Ion Trap |
| Year | Products |
1999 |
Esquire-LC |
| Author | |
Berggard, T.(1,4), Cohen, A.(2), Persson, P.(2), Lindqvist, A.(1), Cedervall, T.(1), Silow, M.(3), Thogersen, I.B.(4), Jonsson, J.-A.(2), Enghild, J.J.(4), Akerstrom, B.(*1) |
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| Reference | |
Protein Sci. 8(12) 1999 2611-2620 |
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Abstract |
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Electrospray (ESP) MS and Edman sequencing are used to identify 21 heterogeneous yellow-brown coloured peptides repurified by RP-HPLC from reduced, carboxymethylated and trypsin digested α1-microglobulin (α1m). The peptides are identified by MS/MS and all peptides in a tryptic digest of α1m are analyzed by peptide mapping using ESP LC/MS. Fluorescence spectra are also measured. Infusion1: 50 μl inj., 50% aq. acetonitrile contg. 0.1% trifluoroacetic acid (TFA), 10 μl/min. MS1: Micromass Bio-Q, ESP, pos. mode, m/z up to 1600. Infusion2: samples in 50% aq. MeOH contg. 0.1% trichloroacetic acid, Au-plated glass capillary. MS2: Bruker Esquire LC, ESP, pos. mode. LC: Isco system, 5 μl inj., 15 cm x 1 mm i.d. (Deltabond ODS), 90% aq. acetonitrile contg. 0.025% TFA, 30 μl/min split 1:1 to MS and UV (350 nm). MS3: Micromass Quattro-BQ, ESP, pos. mode, continuum mode, m/z 230-1450, 5 s/scan. MS/MS spectra of the doubly charged ions indicate the presence of chromophores with masses 122-282 Da, assigned to diff. coloured peptides, covalently attached to Lys-92, Lys-118 and Lys-130 of human α1m. The Lys are semiburied in the lipocalin pocket of the protein. There is also an uncoloured lipophilic, hydroxylated and aminated substance noncovalently associated with the chromophores. |
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