Studies on blood plasma proteome were performed by two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) using samples supplied from China and the United States by collaborations of Human Plasma Proteome Project of HUPO. The plasma proteins were separated by isoelectric focusing using capillary type polyacrylamide gel (PAG) with carrier ampholytes pH 3.5-10 in the first dimension and with SDS-PAGE using 15%, 12.5% and 8% gel in the second dimension. The protein spots were identified by peptide mass fingerprinting using electrospray ionization MS/MS (LC/MSD Trap XCT, Agilent). More than 145 protein spots were identified by LC-MS/MS. The 2-DE patterns showed that the samples stored at 4°C for 3 days lost inter-alpha-trypsin inhibitor heavy chain 4 (ITIH-4).

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We recently demonstrated glycation of monoclonal IgA and the presence of IgA-albumin complexes, but the mechanism of IgA-albumin complex formation was not clear. We isolated the IgA-albumin complexes from 5 IgA type M-proteinemia patients’ sera. To elucidate the mechanism of IgA-albumin complex formation, we performed the dissociation assay of IgA-albumin complexes, the identification of albumin binding sites of monoclonal IgA using immunoelectrophoresis, western blotting and chromatography technologies. In all patients with IgA type M-proteinemia, the IgA-albumin complexes were dissociated by treated with 2-mercaptoethanol (2-ME), but not by treated with a strong acid as acetic acid or NaCl of high concentrations. Moreover, when the purified monoclonal IgA containing IgA-albumin complexes was digested with the IgA protease from Neisseria gonorrhoeae, no macro-albumin was demonstrated. It seems probable that albumin is bound to the monoclonal IgA molecule by covalent disulfide bonds, and that the binding site of albumin is located in near the hinge region of IgA molecule and involve the free SH group thought to be present in the α-chain.

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Wrong:Kayo Yokomizo¹⁾²⁾,Sachiko Maruya²⁾,Nobuo Hiratsuka²⁾ and Kiyoko Shiba²⁾
Right:Kayo Yokomizo¹⁾²⁾,Sachiko Maruya¹⁾,Nobuo Hiratsuka¹⁾ and Kiyoko Shiba¹⁾

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