SUNY at Albany
June 19-23, 2001
Structure Analysis of Serum Amyloid A Isoforms by Limited Proteolysis and Mass Spectrometry
Amyloid fibril formation is the pathologic hallmark of amyloid diseases, which are associated with the self-assembly of otherwise soluble proteins into toxic insoluble cross-beta fibrils. We are using several murine serum amyloid A (SAA) isoforms as a model system to investigate the molecular basis of amyloid formation. In mice, the SAA2 protein is amyloidogenic, whereas the SAA1 and SAAce/j are non-amyloidogenic. These three SAA's are ~12KDa and 103-residues long each, with 9 or less residue differences between them. In this study, the structure and dynamics of the three SAA isoforms were probed by limited proteolysis using trypsin, protease V8, and protease K, and the proteolysis products were analyzed by SDS-PAGE, C4 reverse-phase HPLC and mass spectrometry. The digestion patterns were similar for the three variants, resulting in a protease-resistant fragment of about 5 KDa that corresponds to the central region of SAA. In addition, the proteolytic cleavage of SAA1 and SAA ce/j was slower than for SAA2, indicating either increased flexibility or a slightly different structure for SAA2. The implications of these findings for the mechanism of SAA amyloid fibril formation will be discussed.
Limin Wang, Daniel Moriarty, and Wilfredo Col?n
Department of Chemistry, Rensselaer Polytechnic Institute, Troy, NY, 12180 Ph: 518-276-3027; email: firstname.lastname@example.org