Facile backbone (1H, 15N, 13Ca, and 13C′) assignment of 13C/15N‐labeled proteins using orthogonal projection planes of HNN and HN(C)N experiments and its automation

@article{Kumar2012FacileB,
  title={Facile backbone (1H, 15N, 13Ca, and 13C′) assignment of 13C/15N‐labeled proteins using orthogonal projection planes of HNN and HN(C)N experiments and its automation},
  author={Dinesh Kumar and Aditi N. Borkar and Ramakrishna V. Hosur},
  journal={Magnetic Resonance in Chemistry},
  year={2012},
  volume={50}
}
Recently, we introduced an efficient high‐throughput protocol for backbone assignment of small folded proteins based on two‐dimensional (2D) projections of HN(C)N suite of experiments and its automation [Borkar et al., J. Biomol. NMR 2011, 50(3), 285–297]. This strategy provides complete sequence‐specific assignment of backbone (1H, 15N, 13Cα, and 13C′) resonances in less than a day; thus, it has great implications for high‐throughput structural proteomics. However, in cases when such small… 
Direct Sequential Hit Strategy for Unambiguous and Accurate Backbone Assignment of 13C/15N Labeled Proteins
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An efficient strategy -named here as strategy— has been presented for unambiguous and accurate backbone assignment of 13C/15N labeled proteins using an intrinsically unfolded 164-residue protein named DLC1 binding domain of nNOS.
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The PFBD strategy, because of its speed and simplicity, will be very valuable in Biomolecular NMR community for high-throughput structural proteomics of small folded proteins of MW < 10–12 kDa, the regime where NMR is generally preferred over X-ray crystallography.
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It is believed that the method and protocol proposed here can be used for routine high‐throughput structural studies of proteins and is demonstrated using alpha‐helical bovine apo calbindin‐D9k P43M mutant (75 aa) protein.
Advances in Proteomic Methods
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SummaryA computer algorithm that determines the 1HN, 15N, 13Cα, 1Hα, 13Cβ chemical-shift assignments of protein residues with minimal human intervention is described. The algorithm is implemented as
Automated Resonance Assignment of Proteins: 6 DAPSY-NMR
TLDR
The results demonstrate that APSY-NMR spectroscopy in combination with a suitable assignment algorithm can provide fully automated sequence-specific backbone assignments of small proteins.
Improved 3D triple resonance experiments, HNN and HN(C)N, for HN and 15N sequential correlations in (13C, 15N) labeled proteins: Application to unfolded proteins
Two triple resonance experiments, HNN and HN(C)N, are presented which correlate HN and 15N resonances sequentially along the polypeptide chain of a doubly (13C, 15N) labeled protein. These
AUTOBA: Automation of backbone assignment from HN(C)N suite of experiments
TLDR
An efficient algorithm named as AUTOBA (Automatic Backbone Assignment) designed to automate the assignment protocol based on HN(C)N suite of experiments, demonstrated with experimental spectra recorded on two small globular proteins and simulated spectra of 27 other proteins using assignment data from the BMRB.
Automated probabilistic method for assigning backbone resonances of (13C,15N)-labeled proteins
TLDR
The automated assignment program was tested using CBCANH and CBCA(CO)NH cross peaks of the two previously assigned proteins, calmodulin and CheA, yielding an assignment in excellent agreement with that obtained by time-consuming, manual methods.
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TLDR
It is demonstrated for four small globular proteins that sequential resonance assignment can be routinely obtained within a few hours, or less, in a highly automated and robust way.
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TLDR
It is shown that the 20 amino acid residues can be grouped into eight distinct categories, each of which is assigned a unique two-digit code that is used to tag individual sets of chemical shifts in the master_list and also to translate the protein primary sequence into an array called pps_array.
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TLDR
This paper describes an efficient approach to rapid resonance assignment that is suitable for both folded and unfolded proteins, making use of the triple-resonance experiments described recently, and believes that the new assignment approach will be of great value for both structural genomics and protein folding research by NMR.
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