caa a22 4500 477136621 CHVBK 20180405111720.0 cr unu---uuuuu 170330e19970201xx s 000 0 eng 10.1023/A:1018658305040 doi (NATIONALLICENCE)springer-10.1023/A:1018658305040 Triple-resonance NOESY-based experiments with improved spectral resolution: Applications to structural characterization of unfolded, partially folded and folded proteins [Elektronische Daten] [Ouwen Zhang, Julie Forman-Kay, David Shortle, Lewis Kay] NMR-based structural studies of macromolecules focus to a large extent on the establishmentof interproton distances within the molecule based on the nuclear Overhauser effect (NOE).Despite the improvements in resolution resulting from multidimensional NMR experiments,the detailed characterization of disordered states of proteins or highly overlapped regions offolded molecules using current NMR methods remains challenging. A suite of triple-resonanceNOESY-type pulse schemes is presented which require uniform 15N and 13C labeling andmake use of the chemical shift dispersion of backbone 15N and 13C′ (carbonyl)resonances to increase the spectral resolution. In particular, for the case of partially folded andunfolded proteins, the experiments exploit the fact that the dispersion of 15N and 13C′resonances is comparable to that observed in folded states. Ambiguities that arise in theassignment of NOEs as a result of the severe chemical shift degeneracy in 1H and aliphatic13C nuclei are resolved, therefore, by recording the chemical shifts of 15N or 13C′either before or after the NOE mixing period. Applications of these methods to the study ofthe unfolded state of the N-terminal SH3 domain of drk (drkN SH3) and a partially foldedlarge fragment of staphylococcal nuclease (SNase), Δ131Δ, are presented. Inaddition, an application to folded SNase in complex with the ligands thymidine3′,5′-bisphosphate (pdTp) and Ca2+ is illustrated which allows the assignmentof NOEs between degenerate Hα protons or protons resonating close to water. Kluwer Academic Publishers, 1997 Heteronuclear, multidimensional NMR nationallicence NOE nationallicence Unfolded or partially folded proteins nationallicence Zhang Ouwen Protein Engineering Network Centres of Excellence and Departments of Medical Genetics, Biochemistry and Chemistry, University of Toronto, M5S 1A8, Toronto, ON, Canada aut Forman-Kay Julie Biochemistry Research Division, Hospital for Sick Children, 555 University Avenue, M5G 1X8, Toronto, ON, Canada aut Shortle David Department of Biological Chemistry, Johns Hopkins University School of Medicine, 21205, Baltimore, MD, U.S.A aut Kay Lewis Protein Engineering Network Centres of Excellence and Departments of Medical Genetics, Biochemistry and Chemistry, University of Toronto, M5S 1A8, Toronto, ON, Canada aut Journal of Biomolecular NMR Kluwer Academic Publishers 9/2(1997-02-01), 181-200 0925-2738 9:2<181 1997 9 10858 https://doi.org/10.1023/A:1018658305040 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1023/A:1018658305040 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Zhang Ouwen Protein Engineering Network Centres of Excellence and Departments of Medical Genetics, Biochemistry and Chemistry, University of Toronto, M5S 1A8, Toronto, ON, Canada aut NATIONALLICENCE 700 1- Forman-Kay Julie Biochemistry Research Division, Hospital for Sick Children, 555 University Avenue, M5G 1X8, Toronto, ON, Canada aut NATIONALLICENCE 700 1- Shortle David Department of Biological Chemistry, Johns Hopkins University School of Medicine, 21205, Baltimore, MD, U.S.A aut NATIONALLICENCE 700 1- Kay Lewis Protein Engineering Network Centres of Excellence and Departments of Medical Genetics, Biochemistry and Chemistry, University of Toronto, M5S 1A8, Toronto, ON, Canada aut NATIONALLICENCE 773 0- Journal of Biomolecular NMR Kluwer Academic Publishers 9/2(1997-02-01), 181-200 0925-2738 9:2<181 1997 9 10858 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer