caa a22 4500 445325909 CHVBK 20180317142720.0 cr unu---uuuuu 170323e20110801xx s 000 0 eng 10.1007/s10858-011-9526-0 doi (NATIONALLICENCE)springer-10.1007/s10858-011-9526-0 A segmental labeling strategy for unambiguous determination of domain-domain interactions of large multi-domain proteins [Elektronische Daten] [Jianglei Chen, Jianjun Wang] NMR structural determination of large multi-domain proteins is a challenging task due to significant spectral overlap with a particular difficulty in unambiguous identification of domain-domain interactions. Segmental labeling is a NMR strategy that allows for isotopically labeling one domain and leaves the other domain unlabeled. This significantly simplifies spectral overlaps and allows for quick identification of domain-domain interaction. Here, a novel segmental labeling strategy is presented for detection of inter-domain NOEs. To identify domain-domain interactions in human apolipoprotein E (apoE), a multi-domain, 299-residues α-helical protein, on-column expressed protein ligation was utilized to generate a segmental-labeled apoE samples in which the N-terminal (NT-) domain was 2H(99%)/15N-labeled whereas the C-terminal (CT-) domain was either 15N- or 15N/13C-labeled. 3-D 15N-edited NOESY spectra of these segmental-labeled apoE samples allow for direct observation of the inter-domain NOEs between the backbone amide protons of the NT-domain and the aliphatic protons of the CT-domain. This straightforward approach permits unambiguous identification of 78 inter-domain NOEs, enabling accurate definition of the relative positions of both the NT- and the CT-domains and determination of the NMR structure of apoE. Springer Science+Business Media B.V., 2011 NMR nationallicence Segmental labeling nationallicence NOESY nationallicence NOE nationallicence Inter-domain interaction nationallicence apoE3 nationallicence Chen Jianglei Department of Biochemistry and Molecular Biology, School of Medicine, Wayne State University, 48201, Detroit, MI, USA aut Wang Jianjun Department of Biochemistry and Molecular Biology, School of Medicine, Wayne State University, 48201, Detroit, MI, USA aut Journal of Biomolecular NMR Springer Netherlands 50/4(2011-08-01), 403-410 0925-2738 50:4<403 2011 50 10858 https://doi.org/10.1007/s10858-011-9526-0 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s10858-011-9526-0 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Chen Jianglei Department of Biochemistry and Molecular Biology, School of Medicine, Wayne State University, 48201, Detroit, MI, USA aut NATIONALLICENCE 700 1- Wang Jianjun Department of Biochemistry and Molecular Biology, School of Medicine, Wayne State University, 48201, Detroit, MI, USA aut NATIONALLICENCE 773 0- Journal of Biomolecular NMR Springer Netherlands 50/4(2011-08-01), 403-410 0925-2738 50:4<403 2011 50 10858 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer