The regions of the sequence most exposed to the solvent within the amyloidogenic state of a protein initiate the aggregation process
| Autor: | Giulia Calloni, Fabrizio Chiti, Barbara Lelj Garolla di Bard, Piero Pucci, Gianpietro Ramponi, Maria Chiara Monti, Angela Amoresano |
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| Přispěvatelé: | Monti, Maria, B. L., GAROLLA DI BARD, G., Calloni, F., Chiti, Amoresano, Angela, G., Ramponi, Pucci, Pietro |
| Rok vydání: | 2004 |
| Předmět: |
Amyloid
Protein Folding Spectrometry Mass Electrospray Ionization Protein Conformation Protein design Protein aggregation Acylphosphatase Structural Biology Native state Humans Amino Acid Sequence Molecular Biology mass spectrometry acylphosphatase Chemistry C-terminus Circular Dichroism Proteolytic enzymes aggregation Protein engineering Acid Anhydride Hydrolases limited proteolysi Biochemistry Solvents Peptide Hydrolases |
| Zdroj: | Journal of molecular biology. 336(1) |
| ISSN: | 0022-2836 |
| Popis: | Formation of misfolded aggregates is an essential part of what proteins can do. The process of protein aggregation is central to many human diseases and any aggregating event needs to be prevented within a cell and in protein design. In order to aggregate, a protein needs to unfold its native state, at least partially. The conformational state that is prone to aggregate is difficult to study, due to its aggregating potential and heterogeneous nature. Here, we use a systematic approach of limited proteolysis, in combination with electrospray ionisation mass spectrometry, to investigate the regions that are most flexible and solvent-exposed within the native, ligand-bound and amyloidogenic states of muscle acylphosphatase (AcP), a protein previously shown to form amyloid fibrils in the presence of trifluoroethanol. Seven proteases with different degrees of specificity have been used for this purpose. Following exposure to the aggregating conditions, a number of sites along the sequence of AcP become susceptible to proteolytic digestion. The pattern of proteolytic cleavages obtained under these conditions is considerably different from that of the native and ligand-bound conformations and includes a portion within the N-terminal tail of the protein (residues 6-7), the region of the sequence 18-23 and the position 94 near the C terminus. There is a significant overlap between the regions of the sequence found to be solvent-exposed from the present study and those previously identified to be critical in the rate-determining steps of aggregation from protein engineering approaches. This indicates that a considerable degree of solvent exposure is a feature of the portions of a protein that initiate the process of aggregation. |
| Databáze: | OpenAIRE |
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