期刊
JOURNAL OF MAGNETIC RESONANCE
卷 338, 期 -, 页码 -出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.jmr.2022.107195
关键词
NMR structure; Protein splicing; Halophilic inteins; Segmental isotopic labeling; Conditional protein splicing; Protein engineering
资金
- TEKES [1311/31/2014]
- Academy of Finland [131413, 137995, 277335]
- TaNeDS Europe 2017 grant program from Daiichi Sankyo Co., Ltd
- Novo Nordisk Foundation [NNF17OC0027550]
- Academy of Finland (AKA) [131413, 131413] Funding Source: Academy of Finland (AKA)
Protein trans-splicing catalyzed by split inteins is used to simplify NMR signals by segmental isotopic labeling. We designed a salt-inducible intein with lower salt concentration requirements compared to naturally occurring halo-obligate inteins. The NMR solution structure of the engineered intein showed an identical three-dimensional structure to the original one, albeit it unfolds without salts. The stabilization of the active folded conformation may involve complex interactions, including solvation energy and interactions with water, ions, co-solutes, and protein polypeptide chains.
Protein trans-splicing catalyzed by split inteins has been used for segmental isotopic labeling of proteins for alleviating the complexity of NMR signals. Whereas inteins spontaneously trigger protein splicing upon protein folding, inteins from extremely halophilic organisms require a high salinity condition to induce protein splicing. We designed and created a salt-inducible intein from the widely used DnaE intein from Nostoc punctiforme by introducing 29 mutations, which required a lower salt concentration than naturally occurring halo-obligate inteins. We determined the NMR solution structure of the engineered salt-inducible DnaE intein in 2 M NaCl, showing the essentially identical three-dimensional structure to the original one, albeit it unfolds without salts. The NMR structure of a halo-obligate intein under high salinity suggests that the stabilization of the active folded conformation is not a mere result of various intramolecular interactions but the subtle energy balance from the complex interactions, including the solvation energy, which involve waters, ions, co-solutes, and protein polypeptide chains. (C) 2022 The Authors. Published by Elsevier Inc.
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