Applications for measuring scalar and residual dipolar couplings in proteins
1University of Oulu, Faculty of Science, Department of Chemistry
|Online Access:||PDF Full Text (PDF, 1.4 MB)|
|Persistent link:|| http://urn.fi/urn:isbn:9514258223
|Publish Date:|| 2000-11-03
|Thesis type:||Doctoral Dissertation
|Defence Note:||Academic Dissertation to be presented with the assent of the Faculty of Science, University of Oulu, for public discussion in Raahensali (Auditorium L10), Linnanmaa, on November 25th, 2000, at 12 noon.
Professor Matti Punkkinen
Professor Pekka Pyykkö
Nuclear magnetic resonance spectroscopic structure determination of proteins has been under rapid development during the last decade. The size limitation impeding structural studies of biological macromolecules in solution has increased from 10 kDa to 30 kDa thanks to exploitation of 15N/13C enrichment. Perdeuteration of non-exchangeable protons has pushed this limit even further, allowing backbone resonance assignment of 40 to 50 kDa proteins. Most recently, transverse relaxation optimized spectroscopy (TROSY) has been demonstrated to lengthen 15N and 1HN spin transverse relaxation times significantly, especially in large perdeuterated proteins, thus extending the size limit beyond 100 kDa systems. However, determination of structurally important nuclear Overhauser enhancements (NOE) suffers from perdeuteration, due to the lower density of proton spins available, eventually leading to imprecise protein structures. Very recently, residual dipolar couplings have been used to supplement NOE information, enabling accurate molecular structures to also be obtained with perdeuterated proteins. This thesis focuses on the measurement of the structurally important 3J-coupling between 1HN and 1Hα spins, and determination of residual dipolar couplings by utilizing the novel spin-state-selective subspectral editing together with the TROSY methodology. This approach allows precise measurement of a large number of dipolar couplings in larger protonated or perdeuterated proteins.
Acta Universitatis Ouluensis. A, Scientiae rerum naturalium
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