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J Biomol NMR. 2011 Jan;49(1):3-7. doi: 10.1007/s10858-010-9464-2. Epub 2010 Dec 29.

Observing selected domains in multi-domain proteins via sortase-mediated ligation and NMR spectroscopy.

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  • 1Department of Chemistry, University of Cincinnati, Cincinnati 45221-0172, USA.

Abstract

NMR spectroscopy has distinct advantages for providing insight into protein structures, but faces significant resolution challenges as protein size increases. To alleviate such resonance overlap issues, the ability to produce segmentally labeled proteins is beneficial. Here we show that the S. aureus transpeptidase sortase A can be used to catalyze the ligation of two separately expressed domains of the same protein, MecA (B. subtilis). The yield of purified, segmentally labeled MecA protein conjugate is approximately 40%. The resultant HSQC spectrum obtained from this domain-labeled conjugate demonstrates successful application of sortase A for segmental labeling of multi-domain proteins for solution NMR study.

PMID:
21188472
[PubMed - indexed for MEDLINE]
PMCID:
PMC3111449
Free PMC Article

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