Characterization of high molecular weight multimeric states of human haptoglobin and hemoglobin-based oxygen carriers by high-mass MALDI MS

摘要

High-mass MALDI-TOF mass spectrometry (MS) is a novel analytical approach to study large biomolecules and their interactions. It is a powerful alternative method to gel electrophoresis (GE) and size exclusion chromatography (SEC) for obtaining information on the molecular weights of macromolecules and for determining pro_tein complexes. The precision of mass measurements (mass accuracy), high sensitiv_ity, speed of the analysis, and tolerance toward sample heterogeneity are the major features of this MS-based approach. Remarkably, MS provides direct stoichiometric information of macromolecular protein complexes, when noncovalent interactions are stabilized during desorption ionization by use of chemical cross-linking reagents. In this study, high-mass MALDI-TOF MS was applied to characterize the multimeric state of the human plasma protein haptoglobin (Hp), which is in the mass range of 150 -300 kDa. Also, higher order structures of hemoglobin-based oxy_gen carriers (HBOCs) and their interactions with human haptoglobin were analyzed. These investigations are of clinical importance and contribute to the overall under_standing of specific toxicity and clearance of HBOCs.