NMR - Publications

The successful simulation of proteins by molecular dynamics (MD) critically depends on the accuracy of the applied force field. Here, we modify the AMBER-family ff99SBnmr2 force field through improvements to the side-chain χ(1) dihedral angle potentials in a residue-specific manner using conformational dihedral angle distributions from an experimental coil library as targets. Based on significant deviations observed for the parent force field with respect to the coil library, the χ(1) dihedral...

We describe the preparation, assembly, recognition characteristics, and bioactivity of dendritic basket 6^(12-). This novel cavitand has a deep aromatic pocket with three (S)-glutamic acid dendrons at the rim to amplify water solubility and prevent self-association. ¹H NMR spectroscopy, calorimetry (ITC), and mass spectrometry (ESI-MS) measurements validate the formation of an inclusion complex between 6^(12-) and anticancer drug methotrexate (MTX^(2-)) in water (K(d)=9.2 μM). To identify the...

The field of metabolomics, which is quintessential in today's omics research, involves the large-scale detection, identification, and quantification of small-molecule metabolites in a wide range of biological samples. Nuclear magnetic resonance spectroscopy (NMR) has emerged as a powerful tool for metabolomics due to its high resolution, reproducibility, and exceptional quantitative nature. One of the key bottlenecks of metabolomics studies, however, remains the accurate and automated analysis...

Transforming growth factor (TGF)-β1 is a multifunctional protein that is essential in many cellular processes that include fibrosis, inflammation, chondrogenesis, and cartilage repair. In particular, cartilage repair is important to avoid physical disability since this tissue does not have the inherent capacity to regenerate beyond full development. We report here on supramolecular coassemblies of two peptide amphiphile molecules, one containing a TGF-β1 mimetic peptide, and another which is one...

Despite rapid progress in metabolomics research, a major bottleneck is the large number of metabolites whose chemical structures are unknown or whose spectra have not been deposited in metabolomics databases. Nuclear magnetic resonance (NMR) spectroscopy has a long history of elucidating chemical structures from experimentally measured ¹H and ^(13)C chemical shifts. One approach to characterizing the chemical structures of an unknown metabolite is to predict the ¹H and ^(13)C chemical shifts of...

Human K-Ras protein, which is a member of the GTPase Ras family, hydrolyzes GTP to GDP and concomitantly converts from its active to its inactive state. It is a key oncoprotein, because several mutations, particularly those at residue position 12, occur with a high frequency in a wide range of human cancers. The K-Ras protein is therefore an important target for developing therapeutic anti-cancer agents. In this work we report the almost complete sequence-specific resonance assignments of...

The quantitative deconvolution of 1D-NMR spectra into individual resonances or peaks is a key step in many modern NMR workflows as it critically affects downstream analysis and interpretation. Depending on the complexity of the NMR spectrum, spectral deconvolution can be a notable challenge. Based on the recent deep neural network DEEP Picker and Voigt Fitter for 2D NMR spectral deconvolution, we present here an accurate, fully automated solution for 1D-NMR spectral analysis, including peak...

We describe the preparation, dynamic, assembly characteristics of vase-shaped basket 1^(3-) along with its ability to form an inclusion complex with anticancer drug mitoxantrone in abiotic and biotic systems. This novel cavitand has a deep nonpolar pocket consisting of three naphthalimide sides fused to a bicyclic platform at the bottom while carrying polar glycines at the top. The results of ¹ H Nuclear Magnetic Resonance (NMR), ¹ H NMR Chemical Exchange Saturation Transfer (CEST), Calorimetry,...

An NMR NOAH-supersequence is presented consisting of five CEST experiments for studying protein backbone and side-chain dynamics by ^(15)N-CEST, carbonyl-^(13)CO-CEST, aromatic-^(13)C(ar)-CEST, ^(13)C(α)-CEST, and methyl-^(13)C(met)-CEST. The new sequence acquires the data for these experiments in a fraction of the time required for the individual experiments, saving over four days of NMR time per sample.

AlphaFold2 has revolutionized protein structure prediction from amino-acid sequence. In addition to protein structures, high-resolution dynamics information about various protein regions is important for understanding protein function. Although AlphaFold2 has neither been designed nor trained to predict protein dynamics, it is shown here how the information returned by AlphaFold2 can be used to predict dynamic protein regions at the individual residue level. The approach, which is termed cdsAF2,...

Total joint arthroplasty is a common surgical procedure resulting in improved quality of life; however, a leading cause of surgery failure is infection. Periprosthetic joint infections often involve biofilms, making treatment challenging. The metabolic state of pathogens in the joint space and mechanism of their tolerance to antibiotics and host defenses are not well understood. Thus, there is a critical need for increased understanding of the physiological state of pathogens in the joint space...

An NMR supersequence is introduced for the rapid acquisition of ^(15)N-CEST and methyl-^(13)C-CEST experiments in the same pulse sequence for applications to proteins. The high sensitivity and accuracy allows the simultaneous quantitative characterization of backbone and side-chain dynamics on the millisecond timescale ideal for routine screening for alternative protein states.

Highly quantitative metabolomics studies of complex biological mixtures are facilitated by the resolution enhancement afforded by 2D NMR spectra such as 2D ^(13)C-¹H HSQC spectra. Here, we describe a new public web server, COLMARq, for the semi-automated analysis of sets of 2D HSQC spectra of cohorts of samples. The workflow of COLMARq includes automated peak picking using the deep neural network DEEP Picker, quantitative cross-peak volume extraction by numerical fitting using Voigt Fitter, the...

Rapid progress in machine learning offers new opportunities for the automated analysis of multidimensional NMR spectra ranging from protein NMR to metabolomics applications. Most recently, it has been demonstrated how deep neural networks (DNN) designed for spectral peak picking are capable of deconvoluting highly crowded NMR spectra rivaling the facilities of human experts. Superior DNN-based peak picking is one of a series of critical steps during NMR spectral processing, analysis, and...

There is a critical need to accurately diagnose, prevent, and treat biofilms in humans. The biofilm forming P. aeruginosa bacteria can cause acute and chronic infections, which are difficult to treat due to their ability to evade host defenses along with an inherent antibiotic-tolerance. Using an untargeted NMR-based metabolomics approach, we identified statistically significant differences in 52 metabolites between P. aeruginosa grown in the planktonic and lawn biofilm states. Among them, the...

The analysis of nuclear magnetic resonance (NMR) spectra for the comprehensive and unambiguous identification and characterization of peaks is a difficult, but critically important step in all NMR analyses of complex biological molecular systems. Here, we introduce DEEP Picker, a deep neural network (DNN)-based approach for peak picking and spectral deconvolution which semi-automates the analysis of two-dimensional NMR spectra. DEEP Picker includes 8 hidden convolutional layers and was trained...

Amino-acid side-chain properties in proteins are key determinants of protein function. NMR spin relaxation of side chains is an important source of information about local protein dynamics and flexibility. However, traditional solution NMR relaxation methods are most sensitive to sub-nanosecond dynamics lacking information on slower ns-μs time-scale motions. Nanoparticle-assisted NMR spin relaxation (NASR) of methyl-side chains is introduced here as a window into these ns-μs dynamics. NASR...

The sensitivity-enhanced HSQC, as well as HSQC-TOCSY, experiments have been modified for incorporation into NOAH (NMR by Ordered Acquisition using ¹H detection) supersequences, adding diversity for ^(13)C and ^(15)N modules. Importantly, these heteronuclear modules have been specifically tailored to preserve the magnetisation required for subsequent acquisition of other heteronuclear or homonuclear modules in a supersequence. In addition, we present protocols for optimally combining HSQC and...

Two limiting cases of molecular recognition, induced fit (IF) and conformational selection (CS), play a central role in allosteric regulation of natural systems. The IF paradigm states that a substrate "instructs" the host to change its shape after complexation, while CS asserts that a guest "selects" the optimal fit from an ensemble of preexisting host conformations. With no studies that quantitatively address the interplay of two limiting pathways in abiotic systems, we herein and for the...

Sensitivity-improved versions of two-dimensional (2D) ^(13)C-¹H HSQC (heteronuclear single quantum coherence) and HSQC-TOCSY (HSQC-total correlation spectroscopy) NMR experiments optimized for small biological molecules and their complex mixtures encountered in metabolomics are presented that preserve the magnetization of ¹H spins not directly attached to ^(13)C spins. This allows (i) the application of rapid acquisition techniques to substantially shorten measurement time and (ii) their...