Deconvolution of Two-Dimensional NMR Spectra by Fast Maximum Likelihood Reconstruction: Application to Quantitative Metabolomics

Chylla, Roger A.; Hu, Kaifeng; Ellinger, James J.; Markley, John L.

doi:10.1021/ac200536b

Cited by 86 publications

(107 citation statements)

References 21 publications

Supporting

Mentioning

106

Contrasting

Unclassified

Order By: Relevance

“…Salt was added to the samples used for the H/D exchange measurements because it stabilizes the S state of apo-IscU and made it easier to measure the exchange rates. We used fast maximum-likelihood reconstruction as implemented in the Newton software package (37) in analyzing the hydrogen exchange and 2D NMR exchange data. More experimental details are provided in SI Methods.…”

Section: Methodsmentioning

confidence: 99%

Disordered form of the scaffold protein IscU is the substrate for iron-sulfur cluster assembly on cysteine desulfurase

Kim

Tonelli²,

Markley³

2011

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

169

View full text Add to dashboard Cite

The scaffold protein for iron-sulfur cluster assembly, apo-IscU, populates two interconverting conformational states, one disordered (D) and one structured (S) as revealed by extensive NMR assignments. At pH 8 and 25°C, approximately 70% of the protein is S, and the lifetimes of the states are 1.3 s (S) and 0.50 s (D). Zn (II) and Fe(II) each bind and stabilize structured (S-like) states. Single amino acid substitutions at conserved residues were found that shift the equilibrium toward either the S or the D state. Cluster assembly takes place in the complex between IscU and the cysteine desulfurase, IscS, and our NMR studies demonstrate that IscS binds preferentially the D form of apo-IscU. The addition of 10% IscS to IscU was found to greatly increase H/D exchange at protected amides of IscU, to increase the rate of the S → D reaction, and to decrease the rate of the D → S reaction. In the saturated IscU:IscS complex, IscU is largely disordered. In vitro cluster assembly reactions provided evidence for the functional importance of the S ⇆ D equilibrium. IscU variants that favor the S state were found to undergo a lag phase, not observed with the wild type, that delayed cluster assembly; variants that favor the D state were found to assemble less stable clusters at an intermediate rate without the lag. It appears that IscU has evolved to exist in a disordered conformational state that is the initial substrate for the desulfurase and to convert to a structured state that stabilizes the cluster once it is assembled.amino acid sequence effects on protein stability | protein order-disorder transition | two-dimensional exchange spectroscopy | biogenesis of Fe-S clusters I ron-sulfur (Fe-S) clusters, which are among the most ancient and ubiquitous protein prosthetic groups, function in electron transport, enzymatic catalysis, and chemical sensing reactions or as structural units (1). Humans and other higher eukaryotes utilize the ISC (iron-sulfur cluster) system as the essential Fe-S cluster assembly mechanism in mitochondria, and defects in this system have been linked to a large number of human diseases (2). The prokaryotic ISC system has served as a useful model for understanding Fe-S cluster assembly and delivery. The bacterial system utilizes several proteins that have eukaryotic homologs: IscU (scaffold protein), IscS (cysteine desulfurase), HscB (cochaperone), HscA (chaperone), and CyaY (regulation or iron delivery, analog of human frataxin) (1). Interactions among these proteins have been shown to be critical for efficient Fe-S cluster biogenesis (3). The IscU protein acts as a scaffold on which the Fe-S clusters are assembled and from which the clusters are transferred to various apoproteins. IscS is a homodimeric pyridoxyl-5′-phosphatedependent cysteine desulfurase (4). Each IscS subunit binds an IscU molecule and transfers sulfane sulfur generated from the conversion of cysteine to alanine to the cluster ligand cysteines of IscU (5). HscA and HscB, the DnaK-like chaperone and the DnaJ-like cochaperone pro...

show abstract

Section: Methodsmentioning

confidence: 99%

Disordered form of the scaffold protein IscU is the substrate for iron-sulfur cluster assembly on cysteine desulfurase

Kim

Tonelli²,

Markley³

2011

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

169

View full text Add to dashboard Cite

show abstract

“…The quantification portion of targeted profiling consists of measurement of the intensity of each signal and its profile across the ensemble. Although the "identify" task is usually thought of as a prerequisite to the "quantify" task, a modeling approach such as deconvolution [17][18][19] may quantify signals and modulations of their intensity prior to the signals being assigned to an NMR transition. Fig.…”

Section: Metabolomics Analysis Softwarementioning

confidence: 99%

“…As with ROI-based integrals, ROIbased amplitudes are much more useful for 2D than 1D analysis of complex biological mixtures. The Newton software package for fast maximum likelihood reconstruction of 1D and 2D NMR spectra [19], for example (Fig. 4), makes extensive use of ROI-based amplitudes in performing quantitative analysis of "time-zero" extrapolated 2D HSQC spectra [26].…”

Section: Feature Generationmentioning

confidence: 99%

See 1 more Smart Citation

Databases and Software for NMR-Based Metabolomics

Ellinger¹,

Chylla²,

Ulrich³

et al. 2012

CMB

View full text Add to dashboard Cite

New software and increasingly sophisticated NMR metabolite spectral databases are advancing the unique abilities of NMR spectroscopy to identify and quantify small molecules in solution for studies of metabolite biomarkers and metabolic flux. Public and commercial databases now contain experimental 1D 1 H, 13 C and 2D 1 H-13 C spectra and extracted spectral parameters for over a thousand compounds and theoretical data for thousands more. Public databases containing experimental NMR data from complex metabolic studies are emerging. These databases are providing information vital for the construction and testing of new computational algorithms for NMR-based chemometric and quantitative metabolomics studies. In this review we focus on database and software tools that support a quantitative NMR approach to the analysis of 1D and 2D NMR spectra of complex biological mixtures.

show abstract

“…Two important aspects of spectral deconvolution [29][30][31][32][33] are estimation of the number of signals and proper model function to describe the lineshape. There is overfitting risk to allow a deconvolution algorithm to determine the number of peaks and carry out the fit because increasing the number of peaks will always appear to perfectly fit any spectrum.…”

Section: Nmr Signal Deconvolutionmentioning

confidence: 99%

Quantitative NMR Studies of Multiple Compound Mixtures

2017

Annual Reports on NMR Spectroscopy

View full text Add to dashboard Cite

Deconvolution of Two-Dimensional NMR Spectra by Fast Maximum Likelihood Reconstruction: Application to Quantitative Metabolomics

Cited by 86 publications

References 21 publications

Disordered form of the scaffold protein IscU is the substrate for iron-sulfur cluster assembly on cysteine desulfurase

Disordered form of the scaffold protein IscU is the substrate for iron-sulfur cluster assembly on cysteine desulfurase

Databases and Software for NMR-Based Metabolomics

Quantitative NMR Studies of Multiple Compound Mixtures

Contact Info

Product

Resources

About