title: Direct monitoring of heat-stressed biopolymers with temperature-controlled electrospray ionization mass spectrometry.
abstract: The ability to monitor protein aggregation at the molecular level is critical for progress in many areas of life sciences ranging from understanding mechanisms of amyloidosis and etiology of conformational diseases to development of safe and efficient biopharmaceutical products. Despite the spectacular progress in understanding the mechanisms of protein aggregation in recent years, many aspects of the aggregating proteins behavior remain unclear because of the extreme difficulty in tracking evolution of these notoriously complex and heterogeneous systems. Here, we introduce a mass spectrometry-based methodology that allows the early stages of heat-induced aggregation to be studied by monitoring both conformational changes and formation of oligomers as a function of temperature. The new approach allows biopolymer behavior (both reversible and irreversible processes) to be monitored in a wide temperature range. Validation of the methodology is carried out by comparing temperature profiles of model proteins and nucleic acids deduced from mass spectrometry measurements and differential scanning calorimetry. Application of the methodology to study heat-induced aggregation of human glucocerebrosidase unequivocally links loss of conformational fidelity to formation of soluble oligomers, which serve as precursors to aggregation.
authors.affiliation: Department of Chemistry, University of Massachusetts-Amherst, Amherst, Massachusetts, USA.
authors.forename: Guanbo
authors.initials: G
authors.lastname: Wang
authors.forename: Rinat R
authors.initials: RR
authors.lastname: Abzalimov
authors.forename: Igor A
authors.initials: IA
authors.lastname: Kaltashov
mesh_terms: D001704:Biopolymers / Q000737:chemistry* / Q000378:metabolism; D005962:Glucosylceramidase / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D011487:Protein Conformation; D058767:Protein Unfolding; D021241:Spectrometry, Mass, Electrospray Ionization; D013696:Temperature
chemical_list: D001704:Biopolymers; D005962:Glucosylceramidase