A non-uniform sampling approach enables studies of dilute and unstable proteins

Miljenović, Tomas, Jia, Xinying, Lavrencic, Peter, Kobe, Bostjan and Mobli, Mehdi (2017) A non-uniform sampling approach enables studies of dilute and unstable proteins. Journal of Biomolecular NMR, 1-9. doi:10.1007/s10858-017-0091-z


Author Miljenović, Tomas
Jia, Xinying
Lavrencic, Peter
Kobe, Bostjan
Mobli, Mehdi
Title A non-uniform sampling approach enables studies of dilute and unstable proteins
Journal name Journal of Biomolecular NMR   Check publisher's open access policy
ISSN 1573-5001
0925-2738
Publication date 2017-02-10
Sub-type Article (original research)
DOI 10.1007/s10858-017-0091-z
Open Access Status Not yet assessed
Start page 1
End page 9
Total pages 9
Place of publication Dordrecht, Netherlands
Publisher Springer Netherlands
Language eng
Abstract NMR spectroscopy is a powerful method in structural and functional analysis of macromolecules and has become particularly prevalent in studies of protein structure, function and dynamics. Unique to NMR spectroscopy is the relatively low constraints on sample preparation and the high level of control of sample conditions. Proteins can be studied in a wide range of buffer conditions, e.g. different pHs and variable temperatures, allowing studies of proteins under conditions that are closer to their native environment compared to other structural methods such as X-ray crystallography and electron microscopy. The key disadvantage of NMR is the relatively low sensitivity of the method, requiring either concentrated samples or very lengthy data-acquisition times. Thus, proteins that are unstable or can only be studied in dilute solutions are often considered practically unfeasible for NMR studies. Here, we describe a general method, where non-uniform sampling (NUS) allows for signal averaging to be monitored in an iterative manner, enabling efficient use of spectrometer time, ultimately leading to savings in costs associated with instrument and isotope-labelled protein use. The method requires preparation of multiple aliquots of the protein sample that are flash-frozen and thawed just before acquisition of a short NMR experiments carried out while the protein is stable (12 h in the presented case). Non-uniform sampling enables sufficient resolution to be acquired for each short experiment. Identical NMR datasets are acquired and sensitivity is monitored after each co-added spectrum is reconstructed. The procedure is repeated until sufficient signal-to-noise is obtained. We discuss how maximum entropy reconstruction is used to process the data, and propose a variation on the previously described method of automated parameter selection. We conclude that combining NUS with iterative co-addition is a general approach, and particularly powerful when applied to unstable proteins.
Keyword Jittered sampling
Maximum entropy reconstruction
Non-uniform sampling
Proteins
Targeted acquisition
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

 
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Created: Fri, 17 Feb 2017, 11:15:51 EST by Mrs Louise Nimwegen on behalf of School of Chemistry & Molecular Biosciences