Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation.
The solution structure and stability of N-terminally truncated beta2-microglobulin (deltaN6beta2-m), the major modification in ex vivo fibrils, have been investigated by a variety of biophysical techniques. The results show that deltaN6beta2-m has a free energy of stabilization that is reduced by 2....
| Main Authors: | , , , , , , , , , , , , , , |
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| Format: | Journal article |
| Language: | English |
| Published: |
2000
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| _version_ | 1797053931942051840 |
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| author | Esposito, G Michelutti, R Verdone, G Viglino, P Hernández, H Robinson, C Amoresano, A Dal Piaz, F Monti, M Pucci, P Mangione, P Stoppini, M Merlini, G Ferri, G Bellotti, V |
| author_facet | Esposito, G Michelutti, R Verdone, G Viglino, P Hernández, H Robinson, C Amoresano, A Dal Piaz, F Monti, M Pucci, P Mangione, P Stoppini, M Merlini, G Ferri, G Bellotti, V |
| author_sort | Esposito, G |
| collection | OXFORD |
| description | The solution structure and stability of N-terminally truncated beta2-microglobulin (deltaN6beta2-m), the major modification in ex vivo fibrils, have been investigated by a variety of biophysical techniques. The results show that deltaN6beta2-m has a free energy of stabilization that is reduced by 2.5 kcal/mol compared to the intact protein. Hydrogen exchange of a mixture of the truncated and full-length proteins at microM concentrations at pH 6.5 monitored by electrospray mass spectrometry reveals that deltaN6beta2-m is significantly less protected than its wild-type counterpart. Analysis of deltaN6beta2-m by NMR shows that this loss of protection occurs in beta strands I, III, and part of II. At mM concentration gel filtration analysis shows that deltaN6beta2-m forms a series of oligomers, including trimers and tetramers, and NMR analysis indicates that strand V is involved in intermolecular interactions that stabilize this association. The truncated species of beta2-microglobulin was found to have a higher tendency to self-associate than the intact molecule, and unlike wild-type protein, is able to form amyloid fibrils at physiological pH. Limited proteolysis experiments and analysis by mass spectrometry support the conformational modifications identified by NMR and suggest that deltaN6beta2-m could be a key intermediate of a proteolytic pathway of beta2-microglobulin. Overall, the data suggest that removal of the six residues from the N-terminus of beta2-microglobulin has a major effect on the stability of the overall fold. Part of the tertiary structure is preserved substantially by the disulfide bridge between Cys25 and Cys80, but the pairing between beta-strands far removed from this constrain is greatly perturbed. |
| first_indexed | 2024-03-06T18:50:30Z |
| format | Journal article |
| id | oxford-uuid:100d124f-0aa2-4aa0-a642-0574ed10d8fa |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-06T18:50:30Z |
| publishDate | 2000 |
| record_format | dspace |
| spelling | oxford-uuid:100d124f-0aa2-4aa0-a642-0574ed10d8fa2022-03-26T09:54:24ZRemoval of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:100d124f-0aa2-4aa0-a642-0574ed10d8faEnglishSymplectic Elements at Oxford2000Esposito, GMichelutti, RVerdone, GViglino, PHernández, HRobinson, CAmoresano, ADal Piaz, FMonti, MPucci, PMangione, PStoppini, MMerlini, GFerri, GBellotti, VThe solution structure and stability of N-terminally truncated beta2-microglobulin (deltaN6beta2-m), the major modification in ex vivo fibrils, have been investigated by a variety of biophysical techniques. The results show that deltaN6beta2-m has a free energy of stabilization that is reduced by 2.5 kcal/mol compared to the intact protein. Hydrogen exchange of a mixture of the truncated and full-length proteins at microM concentrations at pH 6.5 monitored by electrospray mass spectrometry reveals that deltaN6beta2-m is significantly less protected than its wild-type counterpart. Analysis of deltaN6beta2-m by NMR shows that this loss of protection occurs in beta strands I, III, and part of II. At mM concentration gel filtration analysis shows that deltaN6beta2-m forms a series of oligomers, including trimers and tetramers, and NMR analysis indicates that strand V is involved in intermolecular interactions that stabilize this association. The truncated species of beta2-microglobulin was found to have a higher tendency to self-associate than the intact molecule, and unlike wild-type protein, is able to form amyloid fibrils at physiological pH. Limited proteolysis experiments and analysis by mass spectrometry support the conformational modifications identified by NMR and suggest that deltaN6beta2-m could be a key intermediate of a proteolytic pathway of beta2-microglobulin. Overall, the data suggest that removal of the six residues from the N-terminus of beta2-microglobulin has a major effect on the stability of the overall fold. Part of the tertiary structure is preserved substantially by the disulfide bridge between Cys25 and Cys80, but the pairing between beta-strands far removed from this constrain is greatly perturbed. |
| spellingShingle | Esposito, G Michelutti, R Verdone, G Viglino, P Hernández, H Robinson, C Amoresano, A Dal Piaz, F Monti, M Pucci, P Mangione, P Stoppini, M Merlini, G Ferri, G Bellotti, V Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation. |
| title | Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation. |
| title_full | Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation. |
| title_fullStr | Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation. |
| title_full_unstemmed | Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation. |
| title_short | Removal of the N-terminal hexapeptide from human beta2-microglobulin facilitates protein aggregation and fibril formation. |
| title_sort | removal of the n terminal hexapeptide from human beta2 microglobulin facilitates protein aggregation and fibril formation |
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