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Segmental isotopic labeling by asparaginyl endopeptidase-mediated protein ligation


Mikula, Kornelia M; Krumwiede, Luisa; Plückthun, Andreas; Iwaï, Hideo (2018). Segmental isotopic labeling by asparaginyl endopeptidase-mediated protein ligation. Journal of Biomolecular NMR, 71(4):225-235.

Abstract

Segmental isotopic labeling can facilitate NMR studies of large proteins, multi-domain proteins, and proteins with repetitive sequences by alleviating NMR signal overlaps. Segmental isotopic labeling also allows us to investigate an individual domain in the context of a full-length protein by NMR. Several established methods are available for segmental isotopic labeling such as intein-mediated ligation, but each has specific requirements and limitations. Here, we report an enzymatic approach using bacterially produced asparagine endopeptidase from Oldenlandia affinis for segmental isotopic labeling of a protein with repetitive sequences, a designed armadillo repeat protein, by overcoming some of the shortcomings of enzymatic ligation for segmental isotopic labeling.

Abstract

Segmental isotopic labeling can facilitate NMR studies of large proteins, multi-domain proteins, and proteins with repetitive sequences by alleviating NMR signal overlaps. Segmental isotopic labeling also allows us to investigate an individual domain in the context of a full-length protein by NMR. Several established methods are available for segmental isotopic labeling such as intein-mediated ligation, but each has specific requirements and limitations. Here, we report an enzymatic approach using bacterially produced asparagine endopeptidase from Oldenlandia affinis for segmental isotopic labeling of a protein with repetitive sequences, a designed armadillo repeat protein, by overcoming some of the shortcomings of enzymatic ligation for segmental isotopic labeling.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Department of Biochemistry
07 Faculty of Science > Department of Biochemistry
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Biochemistry
Physical Sciences > Spectroscopy
Language:English
Date:August 2018
Deposited On:23 Nov 2018 13:47
Last Modified:30 Nov 2023 08:02
Publisher:Springer
ISSN:0925-2738
OA Status:Closed
Free access at:PubMed ID. An embargo period may apply.
Publisher DOI:https://doi.org/10.1007/s10858-018-0175-4
PubMed ID:29536230