Huaping Mo - Academia.edu (original) (raw)

Papers by Huaping Mo

Anal Chem, 2009

Metabolic profiling has received increasing recognition as an indispensable complement to genomic... more Metabolic profiling has received increasing recognition as an indispensable complement to genomics and proteomics for probing biological systems and for clinical applications. (1)H nuclear magnetic resonance (NMR) is widely used in the field but is challenged by spectral complexity and overlap. Improved and simple methods that quantitatively profile a large number of metabolites are sought to make further progress. Here, we demonstrate a simple isotope tagging strategy, in which metabolites with carboxyl groups are chemically tagged with (15)N-ethanolamine and detected using a 2D heteronuclear correlation NMR experiment. This method is capable of detecting over 100 metabolites at concentrations as low as a few micromolar in biological samples, both quantitatively and reproducibly. Carboxyl-containing compounds are found in almost all metabolic pathways, and thus this new approach should find a variety of applications.

Anal Chem, 2010

An increased interest in metabolite profiling is driving the need for improved analytical techniq... more An increased interest in metabolite profiling is driving the need for improved analytical techniques with greater performance for a variety of important applications. Despite their limited sensitivity, nuclear magnetic resonance (NMR) methods are attractive because of their simplicity, reproducibility, quantitative nature, and wide applicability. The use of chemoselective isotopic tags has the potential to advance the application of NMR for analyzing metabolites in complex biofluids by allowing detection of metabolites down to the low micromoalr level with high resolution and specificity. Here, we report a new 13 C-tagging method using 13 C-formic acid that delivers high sensitivity, good quantitation, and excellent resolution for 1 H-13 C 2D NMR profiling of amino metabolites. High reproducibility (coefficient of variation (CV) ) 2%) was observed for metabolites in urine with concentrations down to 10 µM. As amino compounds comprise an important class of metabolites and small molecules of biological roles, this new method therefore should be amenable to a variety of applications.

Alpha-MSH has generally been accepted as the endogenous ligand for melancortin 4 receptors (MC4R)... more Alpha-MSH has generally been accepted as the endogenous ligand for melancortin 4 receptors (MC4R), which plays a major role in energy homeostasis. Targeting MC4R to develop antiobesity agents, many investigators have performed structure-activity relationship (SAR) studies based on α-MSH structure. In this report, we performed a SAR study using human β-MSH(5-22) (DEGPYRMEHFRWGSPPKD, peptide 1) as a lead sequence to develop potent and selective agonists for MC4R and MC3R. The SAR study was begun with a truncation of N-terminus of β-MSH(5-22) together with acetylation of the N-terminus and amidation of the C-terminus of the peptide. Introduction of a cyclic disulfide constrain and replacement of L-Phe with D-Phe afforded a super potent agonist (peptide 5). Further truncation at the C-terminus generated a small and potent MC4R and MC3R agonist (Ac-YRcyclo[CEHdFRWC]amide , peptide 6), which exhibited no MC5R and greatly reduced MC1R activity. Molecular modeling of Ac-YRcyclo[CEHdFRWC]amide (peptide 6) revealed that Arg2 in the peptide formed a salt bridge with Glu4. Subcutaneous (SC) or intracebroventricular (ICV) administration of peptide 6 in rats showed potent in vivo efficacy as evidenced by its effects in reducing energy balance, increasing fat utilization and decreasing weight gain in both acute and chronic rat metabolic studies.

Nature Structural Biology, 2002

Here we report the structure of acireductone dioxygenase (ARD), the first determined for a new fa... more Here we report the structure of acireductone dioxygenase (ARD), the first determined for a new family of metalloenzymes. ARD represents a branch point in the methionine salvage pathway leading from methylthioadenosine to methionine and has been shown to catalyze different reactions depending on the type of metal ion bound in the active site. The solution structure of nickel-containing ARD (Ni-ARD) was determined using NMR methods. X-ray absorption spectroscopy, assignment of hyperfine shifted NMR resonances and conserved domain homology were used to model the metal-binding site because of the paramagnetism of the bound Ni2+. Although there is no structure in the Protein Data Bank within 3 A r.m.s deviation of that of Ni-ARD, the enzyme active site is located in a conserved double-stranded b-helix domain. Furthermore, the proposed Ni-ARD active site shows significant post-facto structural homology to the active sites of several metalloenzymes in the cupin superfamily.

The Analyst, 2012

A convenient and fast method for quantifying urea in biofluids is demonstrated using NMR analysis... more A convenient and fast method for quantifying urea in biofluids is demonstrated using NMR analysis and the solvent water signal as a concentration reference. The urea concentration can be accurately determined with errors less than 3% between 1 mM and 50 mM, and less than 2% above 50 mM in urine and serum. The method is promising for various applications with advantages of simplicity, high accuracy, and fast non-destructive detection. With an ability to measure other metabolites simultaneously, this NMR method is also likely to find applications in metabolic profiling and system biology.

Proceedings of the National Academy of Sciences, 2001

The downstream prion-like protein (doppel, or Dpl) is a paralog of the cellular prion protein, Pr... more The downstream prion-like protein (doppel, or Dpl) is a paralog of the cellular prion protein, PrP(C). The two proteins have approximately 25% sequence identity, but seem to have distinct physiologic roles. Unlike PrP(C), Dpl does not support prion replication; instead, overexpression of Dpl in the brain seems to cause a completely different neurodegenerative disease. We report the solution structure of a fragment of recombinant mouse Dpl (residues 26-157) containing a globular domain with three helices and a small amount of beta-structure. Overall, the topology of Dpl is very similar to that of PrP(C). Significant differences include a marked kink in one of the helices in Dpl, and a different orientation of the two short beta-strands. Although the two proteins most likely arose through duplication of a single ancestral gene, the relationship is now so distant that only the structures retain similarity; the functions have diversified along with the sequence.

MedChemComm, 2013

ABSTRACT The sesquiterpene lactone class of natural products displays a diverse array of biologic... more ABSTRACT The sesquiterpene lactone class of natural products displays a diverse array of biological activities due to the presence of the α-methylene-γ-lactone motif. However, clinical translation of this class has been hampered by poor aqueous solubility and non-selective binding as a Michael acceptor at undesired targets. A prodrug approach has been developed to overcome these problems in which an amine is added into the α-methylene-γ-lactone to mask this group from nucleophiles and increase solubility. The medicinal chemistry of amino-derivatives of the sesquiterpene lactones is described, beginning with synthetic development, moving into pharmacological applications, and finishing with clinical translation.

Magnetic Resonance in Chemistry, 2010

The observed NMR signal size depends on the receiver gain parameter. We propose a receiver gain f... more The observed NMR signal size depends on the receiver gain parameter. We propose a receiver gain function to characterize how much the raw FID is amplified by the receiver as a function of the receiver gain setting. Although the receiver is linear for a fixed gain setting, the actual gain of the receiver may differ from what the gain setting suggests. Nevertheless, for a given receiver, we demonstrate that the receiver gain function can be calibrated. Such a calibration enables accurate comparison of separately acquired NMR signals in quantitative analysis, which frequently requires different receiver gain settings to avoid receiver saturation or achieve optimum sensitivity. The application of receiver gain function, along with the definition of receiving efficiency, allows easy concentration determination by a single internal or external concentration reference.

Magnetic Resonance in Chemistry, 2011

The NMR peak integral is ideally linearly dependent on the sine of excitation angle (θ), which ha... more The NMR peak integral is ideally linearly dependent on the sine of excitation angle (θ), which has provided unsurpassed flexibility in quantitative NMR by allowing the use of a signal of any concentration as the internal concentration reference. Controlling the excitation angle is particularly critical for solvent proton concentration referencing to minimize the negative impact of radiation damping, and to reduce the risk of receiver gain compression. In practice, due to the influence of RF inhomogeneity for any given probe, the observed peak integral is not exactly proportional to sin θ. To evaluate the impact quantitatively, we introduce a RF inhomogeneity factor I(θ) as a function of the nominal pulse excitation angle and propose a simple calibration procedure. Alternatively, I(θ) can be calculated from the probe's RF profile, which can be readily obtained as a gradient image of an aqueous sample. Our results show that without consideration of I(θ), even for a probe with good RF homogeneity, up to 5% error can be introduced due to different excitation pulse angles used for the analyte and the reference. Hence, a simple calibration of I(θ) can eliminate such errors and allow an accurate description of the observed NMR signal's dependence on the excitation angle in quantitative analysis.

Magnetic Resonance in Chemistry, 2010

Modern NMR spectrometers require receivers to work within their linear ranges to maintain high fi... more Modern NMR spectrometers require receivers to work within their linear ranges to maintain high fidelity line shape and peak integration. For better sensitivity, the receiver gain has to be optimized to detect dilute analytes; however, gain compression needs to be avoided. Here, we explore if and how linear receiver performance can be achieved for a couple of representative gain settings on a spectrometer. In the case of slight receiver gain compression, not only will the peak integral be attenuated but a very small line-shape change can also be observed. Hence, we can resort to resonance integration and line-shape analysis for gain compression diagnosis. As such, NMR signals, regardless of their observed amplitude difference in frequency domain, can be accurately compared in quantitative analysis.

Journal of the American Chemical Society, 2003

The crucial step in drug discovery is the identification of a lead compound from a vast chemical ... more The crucial step in drug discovery is the identification of a lead compound from a vast chemical library by any number of screening techniques. NMR-based screening has the advantage of directly detecting binding of a compound to the target. The spectra resulting from these screens can also be very complex and difficult to analyze, making this an inefficient process. We present here a method, RAMPED-UP NMR, (Rapid Analysis and Multiplexing of Experimentally Discriminated Uniquely Labeled Proteins using NMR) which generates simple spectra which are easy to interpret and allows several proteins to be screened simultaneously. In this method, the proteins to be screened are uniquely labeled with one amino acid type. There are several benefits derived from this unique labeling strategy: the spectra are greatly simplified, resonances that are most likely to be affected by binding are the only ones observed, and peaks that yield little or no information upon binding are eliminated, allowing the analysis of multiple proteins easily and simultaneously. We demonstrate the ability of three different proteins to be analyzed simultaneously for binding to two different ligands. This method will have significant impact in the use of NMR spectroscopy for both the lead generation and lead optimization phases of drug discovery by its ability to increase screening throughput and the ability to examine selectivity. To the best of our knowledge, this is the first time in any format that multiple proteins can be screened in one tube.

Journal of the American Chemical Society, 1997

Tetrabutylammonium ion (1) forms tight ion pairs with small anions (Cl -, BH 4 -) in CDCl 3 solut... more Tetrabutylammonium ion (1) forms tight ion pairs with small anions (Cl -, BH 4 -) in CDCl 3 solution. These ion pairs aggregate as a response to increasing solution concentration with little temperature dependence. Maximum aggregate size is approximately four ion pairs, as measured by comparing self-diffusion coefficients of the aggregates with that of an internal nonaggregating standard of the same shape and nominal size, tetrabutylsilane (2). The magnitudes of steady state interionic 1 H{ 1 H} NOEs observed between 1 and the BH 4 -anion in CDCl 3 as a function of temperature in solutions of fixed concentration are well fit to the standard theoretical expression by assuming a single aggregate size that is independent of temperature. A simplified model-free analysis was applied to steady state 15 N{ 1 H} NOE and 15 N T 1 measured at several magnetic field strengths, using 15 N-labeled 1 to obtain estimates for reorientational correlation times for the ion aggregates. A similar analysis of 13 C{ 1 H} NOE and 13 C T 1 gives local effective correlation times for C-H bond vectors of the 1-CH 2 carbon of 1 and order parameters relating the local motion to overall cation motion. Comparison of these correlation times with those obtained from analysis of 29 Si{ 1 H} NOE, 13 C{ 1 H} NOE, and 13 C T 1 for silane 2 provides an estimate of aggregate size which is independent of that obtained by diffusion, with good agreement between the different approaches. JA9723139 log η ) -3.646 + 584.9t -1 + (7.22 × 10 -3 )t -(8.26 × 10 -6 )τ 2

Journal of Polymer Science Part A: Polymer Chemistry, 2011

Journal of Medicinal Chemistry, 2011

The design, synthesis, and biological activity of fluorinated amino-derivatives of the sesquiterp... more The design, synthesis, and biological activity of fluorinated amino-derivatives of the sesquiterpene lactone, parthenolide, are described. A fluorinated aminoparthenolide analogue with biological activity similar to the parent natural product was discovered, and its X-ray structure was obtained. This lead compound was then studied using (19)F NMR in the presence and absence of glutathione to obtain additional mechanism of action data, and it was found that the aminoparthenolide eliminates amine faster in the presence of glutathione than in the absence of glutathione. The exact changes in concentrations of fluorinated compound and amine were quantified by a concentration-reference method using (19)F NMR; a major benefit of applying this strategy is that no deuterated solvents or internal standards are required to obtain accurate concentrations. These mechanistic data with glutathione may contribute to the conversion of the amino-derivative to parthenolide, the active pharmacological agent, in glutathione-rich cancer cells.

Journal of Magnetic Resonance, 2009

Recognizing that the sensitivity of NMR is influenced by factors such as conductance and dielectr... more Recognizing that the sensitivity of NMR is influenced by factors such as conductance and dielectric constant of the sample, we propose the receiving efficiency ℜ to characterize how efficiently the NMR signal can be observed from a unit transverse magnetization in a sample under optimal probe tuning and matching conditions. Conveniently, the relative receiving efficiency can be defined as the ratio of the NMR signal induced by a unit transverse magnetization in a sample of interest and a reference solution. Based on the reciprocal relationship between excitation and observation in NMR, the relative receiving efficiency can be correlated with the 90° pulse length (τ 90 ). In the special case of perfect probe tuning (impedance matched to 50 Ω), ℜ is inversely proportional to τ 90 . Application of the NMR receiving efficiency in quantitative analysis potentially enables a single external concentration reference for almost any sample, eliminating the need to know its exact chemical composition or detailed electromagnetic properties.

Endocrinology, 2005

␣MSH has generally been accepted as the endogenous ligand for melanocortin 4 receptor (MC4R), whi... more ␣MSH has generally been accepted as the endogenous ligand for melanocortin 4 receptor (MC4R), which plays a major role in energy homeostasis. Targeting MC4R to develop antiobesity agents, many investigators have performed a structureactivity relationship (SAR) studies based on ␣MSH structure. In this report, we performed a SAR study using human ␤MSH (5-22) (DEGPYRMEHFRWGSPPKD, peptide 1) as a lead sequence to develop potent and selective agonists for MC4R and MC3R. The SAR study was begun with a truncation of N terminus of ␤MSH (5-22) together with acetylation of the N terminus and amidation of the C terminus of the peptide. Introduction of a cyclic disulfide constrain and replacement of L-Phe with D-Phe afforded a super potent agonist (peptide 5). Furthermore truncation at the C terminus generated a small and potent MC4R and MC3R agonist (Ac-YRcyclo[CEHdFRW-C]amide, peptide 6), which exhibited no MC5R and greatly reduced MC1R activity. Molecular modeling of Ac-YRcyclo-[CEHdFRWC]amide (peptide 6) revealed that Arg2 in the peptide formed a salt bridge with Glu4. Subcutaneous or intracerebroventricular administration of peptide 6 in rats showed potent in vivo efficacy as evidenced by its effects in reducing energy balance, increasing fat use, and decreasing weight gain in both acute and chronic rat metabolic studies. Furthermore, the antiobesity effect by peptide 6 was manifested only in wild-type but not MC4R-deficient mice, indicating that antiobesity effects of the peptide were attributed largely through MC4R but not MC3R agonist activity of the peptide.

Carbohydrate Research, 2012

With BF 3 ÁOEt 2 as the catalyst, the glycosylation of phenols with glycosyl trichloroacetimidate... more With BF 3 ÁOEt 2 as the catalyst, the glycosylation of phenols with glycosyl trichloroacetimidates (or N-phenyl trifluoroacetimidates) bearing 2-O-participating groups leads to the desired 1,2-trans-O-glycosides in generally excellent yields without formation of the 1,2-cis-anomers. However, with TMSOTf as the catalyst, the outcomes of the corresponding phenol O-glycosylation are highly dependent on the nucleophilicity of the phenols; less nucleophilic is the phenol, higher amounts of the 1,2-cis-O-glycoside together with more side-products are generated. 1,2-Orthoesters have been found to be the major products at a low temperature (<À70°C) in all these phenol O-glycosylation reactions, which are transformed into the final products at a higher temperature. BF 3 ÁOEt 2 is an effective catalyst to promote the conversion of 1,2-orthoesters into the corresponding 1,2-trans-O-glycosides. However, the 1,2-orthoesters could be converted into the dioxolenium triflate and glycosyl triflate in the presence of TMSOTf, these intermediates which might be in equilibrium with the glycosyl oxocarbenium related species lead to the final mixture of the a/b-O-glycosides and side-products.

Bioorganic & Medicinal Chemistry Letters, 2013

The biological role of installing a critical exocyclic enone into the structure of the alkaloid, ... more The biological role of installing a critical exocyclic enone into the structure of the alkaloid, (À)-eburnamonine, and characterization of the new chemical reactivity by quantitative NMR without using deuterated solvents are described. This selective modification to a natural product imparts potent anticancer activity as well as bestows chemical reactivity toward nucleophilic thiols, which was measured by quantitative NMR. The synthetic strategy provides an overall conversion of 40%. In the key synthetic step, a modified Peterson olefination was accomplished through the facile release of trifluoroacetate to create the requisite enone in the presence of substantial steric hindrance. j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / b m c l activity in HL-60 (human leukemia) and cytotoxicity in MDA-MB-231 (breast cancer) cells (LC 50 = 14.1 lM); however, the parent

Anal Chem, 2009

Metabolic profiling has received increasing recognition as an indispensable complement to genomic... more Metabolic profiling has received increasing recognition as an indispensable complement to genomics and proteomics for probing biological systems and for clinical applications. (1)H nuclear magnetic resonance (NMR) is widely used in the field but is challenged by spectral complexity and overlap. Improved and simple methods that quantitatively profile a large number of metabolites are sought to make further progress. Here, we demonstrate a simple isotope tagging strategy, in which metabolites with carboxyl groups are chemically tagged with (15)N-ethanolamine and detected using a 2D heteronuclear correlation NMR experiment. This method is capable of detecting over 100 metabolites at concentrations as low as a few micromolar in biological samples, both quantitatively and reproducibly. Carboxyl-containing compounds are found in almost all metabolic pathways, and thus this new approach should find a variety of applications.

Anal Chem, 2010

An increased interest in metabolite profiling is driving the need for improved analytical techniq... more An increased interest in metabolite profiling is driving the need for improved analytical techniques with greater performance for a variety of important applications. Despite their limited sensitivity, nuclear magnetic resonance (NMR) methods are attractive because of their simplicity, reproducibility, quantitative nature, and wide applicability. The use of chemoselective isotopic tags has the potential to advance the application of NMR for analyzing metabolites in complex biofluids by allowing detection of metabolites down to the low micromoalr level with high resolution and specificity. Here, we report a new 13 C-tagging method using 13 C-formic acid that delivers high sensitivity, good quantitation, and excellent resolution for 1 H-13 C 2D NMR profiling of amino metabolites. High reproducibility (coefficient of variation (CV) ) 2%) was observed for metabolites in urine with concentrations down to 10 µM. As amino compounds comprise an important class of metabolites and small molecules of biological roles, this new method therefore should be amenable to a variety of applications.

Alpha-MSH has generally been accepted as the endogenous ligand for melancortin 4 receptors (MC4R)... more Alpha-MSH has generally been accepted as the endogenous ligand for melancortin 4 receptors (MC4R), which plays a major role in energy homeostasis. Targeting MC4R to develop antiobesity agents, many investigators have performed structure-activity relationship (SAR) studies based on α-MSH structure. In this report, we performed a SAR study using human β-MSH(5-22) (DEGPYRMEHFRWGSPPKD, peptide 1) as a lead sequence to develop potent and selective agonists for MC4R and MC3R. The SAR study was begun with a truncation of N-terminus of β-MSH(5-22) together with acetylation of the N-terminus and amidation of the C-terminus of the peptide. Introduction of a cyclic disulfide constrain and replacement of L-Phe with D-Phe afforded a super potent agonist (peptide 5). Further truncation at the C-terminus generated a small and potent MC4R and MC3R agonist (Ac-YRcyclo[CEHdFRWC]amide , peptide 6), which exhibited no MC5R and greatly reduced MC1R activity. Molecular modeling of Ac-YRcyclo[CEHdFRWC]amide (peptide 6) revealed that Arg2 in the peptide formed a salt bridge with Glu4. Subcutaneous (SC) or intracebroventricular (ICV) administration of peptide 6 in rats showed potent in vivo efficacy as evidenced by its effects in reducing energy balance, increasing fat utilization and decreasing weight gain in both acute and chronic rat metabolic studies.

Nature Structural Biology, 2002

Here we report the structure of acireductone dioxygenase (ARD), the first determined for a new fa... more Here we report the structure of acireductone dioxygenase (ARD), the first determined for a new family of metalloenzymes. ARD represents a branch point in the methionine salvage pathway leading from methylthioadenosine to methionine and has been shown to catalyze different reactions depending on the type of metal ion bound in the active site. The solution structure of nickel-containing ARD (Ni-ARD) was determined using NMR methods. X-ray absorption spectroscopy, assignment of hyperfine shifted NMR resonances and conserved domain homology were used to model the metal-binding site because of the paramagnetism of the bound Ni2+. Although there is no structure in the Protein Data Bank within 3 A r.m.s deviation of that of Ni-ARD, the enzyme active site is located in a conserved double-stranded b-helix domain. Furthermore, the proposed Ni-ARD active site shows significant post-facto structural homology to the active sites of several metalloenzymes in the cupin superfamily.

The Analyst, 2012

A convenient and fast method for quantifying urea in biofluids is demonstrated using NMR analysis... more A convenient and fast method for quantifying urea in biofluids is demonstrated using NMR analysis and the solvent water signal as a concentration reference. The urea concentration can be accurately determined with errors less than 3% between 1 mM and 50 mM, and less than 2% above 50 mM in urine and serum. The method is promising for various applications with advantages of simplicity, high accuracy, and fast non-destructive detection. With an ability to measure other metabolites simultaneously, this NMR method is also likely to find applications in metabolic profiling and system biology.

Proceedings of the National Academy of Sciences, 2001

The downstream prion-like protein (doppel, or Dpl) is a paralog of the cellular prion protein, Pr... more The downstream prion-like protein (doppel, or Dpl) is a paralog of the cellular prion protein, PrP(C). The two proteins have approximately 25% sequence identity, but seem to have distinct physiologic roles. Unlike PrP(C), Dpl does not support prion replication; instead, overexpression of Dpl in the brain seems to cause a completely different neurodegenerative disease. We report the solution structure of a fragment of recombinant mouse Dpl (residues 26-157) containing a globular domain with three helices and a small amount of beta-structure. Overall, the topology of Dpl is very similar to that of PrP(C). Significant differences include a marked kink in one of the helices in Dpl, and a different orientation of the two short beta-strands. Although the two proteins most likely arose through duplication of a single ancestral gene, the relationship is now so distant that only the structures retain similarity; the functions have diversified along with the sequence.

MedChemComm, 2013

ABSTRACT The sesquiterpene lactone class of natural products displays a diverse array of biologic... more ABSTRACT The sesquiterpene lactone class of natural products displays a diverse array of biological activities due to the presence of the α-methylene-γ-lactone motif. However, clinical translation of this class has been hampered by poor aqueous solubility and non-selective binding as a Michael acceptor at undesired targets. A prodrug approach has been developed to overcome these problems in which an amine is added into the α-methylene-γ-lactone to mask this group from nucleophiles and increase solubility. The medicinal chemistry of amino-derivatives of the sesquiterpene lactones is described, beginning with synthetic development, moving into pharmacological applications, and finishing with clinical translation.

Magnetic Resonance in Chemistry, 2010

The observed NMR signal size depends on the receiver gain parameter. We propose a receiver gain f... more The observed NMR signal size depends on the receiver gain parameter. We propose a receiver gain function to characterize how much the raw FID is amplified by the receiver as a function of the receiver gain setting. Although the receiver is linear for a fixed gain setting, the actual gain of the receiver may differ from what the gain setting suggests. Nevertheless, for a given receiver, we demonstrate that the receiver gain function can be calibrated. Such a calibration enables accurate comparison of separately acquired NMR signals in quantitative analysis, which frequently requires different receiver gain settings to avoid receiver saturation or achieve optimum sensitivity. The application of receiver gain function, along with the definition of receiving efficiency, allows easy concentration determination by a single internal or external concentration reference.

Magnetic Resonance in Chemistry, 2011

The NMR peak integral is ideally linearly dependent on the sine of excitation angle (θ), which ha... more The NMR peak integral is ideally linearly dependent on the sine of excitation angle (θ), which has provided unsurpassed flexibility in quantitative NMR by allowing the use of a signal of any concentration as the internal concentration reference. Controlling the excitation angle is particularly critical for solvent proton concentration referencing to minimize the negative impact of radiation damping, and to reduce the risk of receiver gain compression. In practice, due to the influence of RF inhomogeneity for any given probe, the observed peak integral is not exactly proportional to sin θ. To evaluate the impact quantitatively, we introduce a RF inhomogeneity factor I(θ) as a function of the nominal pulse excitation angle and propose a simple calibration procedure. Alternatively, I(θ) can be calculated from the probe&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s RF profile, which can be readily obtained as a gradient image of an aqueous sample. Our results show that without consideration of I(θ), even for a probe with good RF homogeneity, up to 5% error can be introduced due to different excitation pulse angles used for the analyte and the reference. Hence, a simple calibration of I(θ) can eliminate such errors and allow an accurate description of the observed NMR signal&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39;s dependence on the excitation angle in quantitative analysis.

Magnetic Resonance in Chemistry, 2010

Modern NMR spectrometers require receivers to work within their linear ranges to maintain high fi... more Modern NMR spectrometers require receivers to work within their linear ranges to maintain high fidelity line shape and peak integration. For better sensitivity, the receiver gain has to be optimized to detect dilute analytes; however, gain compression needs to be avoided. Here, we explore if and how linear receiver performance can be achieved for a couple of representative gain settings on a spectrometer. In the case of slight receiver gain compression, not only will the peak integral be attenuated but a very small line-shape change can also be observed. Hence, we can resort to resonance integration and line-shape analysis for gain compression diagnosis. As such, NMR signals, regardless of their observed amplitude difference in frequency domain, can be accurately compared in quantitative analysis.

Journal of the American Chemical Society, 2003

The crucial step in drug discovery is the identification of a lead compound from a vast chemical ... more The crucial step in drug discovery is the identification of a lead compound from a vast chemical library by any number of screening techniques. NMR-based screening has the advantage of directly detecting binding of a compound to the target. The spectra resulting from these screens can also be very complex and difficult to analyze, making this an inefficient process. We present here a method, RAMPED-UP NMR, (Rapid Analysis and Multiplexing of Experimentally Discriminated Uniquely Labeled Proteins using NMR) which generates simple spectra which are easy to interpret and allows several proteins to be screened simultaneously. In this method, the proteins to be screened are uniquely labeled with one amino acid type. There are several benefits derived from this unique labeling strategy: the spectra are greatly simplified, resonances that are most likely to be affected by binding are the only ones observed, and peaks that yield little or no information upon binding are eliminated, allowing the analysis of multiple proteins easily and simultaneously. We demonstrate the ability of three different proteins to be analyzed simultaneously for binding to two different ligands. This method will have significant impact in the use of NMR spectroscopy for both the lead generation and lead optimization phases of drug discovery by its ability to increase screening throughput and the ability to examine selectivity. To the best of our knowledge, this is the first time in any format that multiple proteins can be screened in one tube.

Journal of the American Chemical Society, 1997

Tetrabutylammonium ion (1) forms tight ion pairs with small anions (Cl -, BH 4 -) in CDCl 3 solut... more Tetrabutylammonium ion (1) forms tight ion pairs with small anions (Cl -, BH 4 -) in CDCl 3 solution. These ion pairs aggregate as a response to increasing solution concentration with little temperature dependence. Maximum aggregate size is approximately four ion pairs, as measured by comparing self-diffusion coefficients of the aggregates with that of an internal nonaggregating standard of the same shape and nominal size, tetrabutylsilane (2). The magnitudes of steady state interionic 1 H{ 1 H} NOEs observed between 1 and the BH 4 -anion in CDCl 3 as a function of temperature in solutions of fixed concentration are well fit to the standard theoretical expression by assuming a single aggregate size that is independent of temperature. A simplified model-free analysis was applied to steady state 15 N{ 1 H} NOE and 15 N T 1 measured at several magnetic field strengths, using 15 N-labeled 1 to obtain estimates for reorientational correlation times for the ion aggregates. A similar analysis of 13 C{ 1 H} NOE and 13 C T 1 gives local effective correlation times for C-H bond vectors of the 1-CH 2 carbon of 1 and order parameters relating the local motion to overall cation motion. Comparison of these correlation times with those obtained from analysis of 29 Si{ 1 H} NOE, 13 C{ 1 H} NOE, and 13 C T 1 for silane 2 provides an estimate of aggregate size which is independent of that obtained by diffusion, with good agreement between the different approaches. JA9723139 log η ) -3.646 + 584.9t -1 + (7.22 × 10 -3 )t -(8.26 × 10 -6 )τ 2

Journal of Polymer Science Part A: Polymer Chemistry, 2011

Journal of Medicinal Chemistry, 2011

The design, synthesis, and biological activity of fluorinated amino-derivatives of the sesquiterp... more The design, synthesis, and biological activity of fluorinated amino-derivatives of the sesquiterpene lactone, parthenolide, are described. A fluorinated aminoparthenolide analogue with biological activity similar to the parent natural product was discovered, and its X-ray structure was obtained. This lead compound was then studied using (19)F NMR in the presence and absence of glutathione to obtain additional mechanism of action data, and it was found that the aminoparthenolide eliminates amine faster in the presence of glutathione than in the absence of glutathione. The exact changes in concentrations of fluorinated compound and amine were quantified by a concentration-reference method using (19)F NMR; a major benefit of applying this strategy is that no deuterated solvents or internal standards are required to obtain accurate concentrations. These mechanistic data with glutathione may contribute to the conversion of the amino-derivative to parthenolide, the active pharmacological agent, in glutathione-rich cancer cells.

Journal of Magnetic Resonance, 2009

Recognizing that the sensitivity of NMR is influenced by factors such as conductance and dielectr... more Recognizing that the sensitivity of NMR is influenced by factors such as conductance and dielectric constant of the sample, we propose the receiving efficiency ℜ to characterize how efficiently the NMR signal can be observed from a unit transverse magnetization in a sample under optimal probe tuning and matching conditions. Conveniently, the relative receiving efficiency can be defined as the ratio of the NMR signal induced by a unit transverse magnetization in a sample of interest and a reference solution. Based on the reciprocal relationship between excitation and observation in NMR, the relative receiving efficiency can be correlated with the 90° pulse length (τ 90 ). In the special case of perfect probe tuning (impedance matched to 50 Ω), ℜ is inversely proportional to τ 90 . Application of the NMR receiving efficiency in quantitative analysis potentially enables a single external concentration reference for almost any sample, eliminating the need to know its exact chemical composition or detailed electromagnetic properties.

Endocrinology, 2005

␣MSH has generally been accepted as the endogenous ligand for melanocortin 4 receptor (MC4R), whi... more ␣MSH has generally been accepted as the endogenous ligand for melanocortin 4 receptor (MC4R), which plays a major role in energy homeostasis. Targeting MC4R to develop antiobesity agents, many investigators have performed a structureactivity relationship (SAR) studies based on ␣MSH structure. In this report, we performed a SAR study using human ␤MSH (5-22) (DEGPYRMEHFRWGSPPKD, peptide 1) as a lead sequence to develop potent and selective agonists for MC4R and MC3R. The SAR study was begun with a truncation of N terminus of ␤MSH (5-22) together with acetylation of the N terminus and amidation of the C terminus of the peptide. Introduction of a cyclic disulfide constrain and replacement of L-Phe with D-Phe afforded a super potent agonist (peptide 5). Furthermore truncation at the C terminus generated a small and potent MC4R and MC3R agonist (Ac-YRcyclo[CEHdFRW-C]amide, peptide 6), which exhibited no MC5R and greatly reduced MC1R activity. Molecular modeling of Ac-YRcyclo-[CEHdFRWC]amide (peptide 6) revealed that Arg2 in the peptide formed a salt bridge with Glu4. Subcutaneous or intracerebroventricular administration of peptide 6 in rats showed potent in vivo efficacy as evidenced by its effects in reducing energy balance, increasing fat use, and decreasing weight gain in both acute and chronic rat metabolic studies. Furthermore, the antiobesity effect by peptide 6 was manifested only in wild-type but not MC4R-deficient mice, indicating that antiobesity effects of the peptide were attributed largely through MC4R but not MC3R agonist activity of the peptide.

Carbohydrate Research, 2012

With BF 3 ÁOEt 2 as the catalyst, the glycosylation of phenols with glycosyl trichloroacetimidate... more With BF 3 ÁOEt 2 as the catalyst, the glycosylation of phenols with glycosyl trichloroacetimidates (or N-phenyl trifluoroacetimidates) bearing 2-O-participating groups leads to the desired 1,2-trans-O-glycosides in generally excellent yields without formation of the 1,2-cis-anomers. However, with TMSOTf as the catalyst, the outcomes of the corresponding phenol O-glycosylation are highly dependent on the nucleophilicity of the phenols; less nucleophilic is the phenol, higher amounts of the 1,2-cis-O-glycoside together with more side-products are generated. 1,2-Orthoesters have been found to be the major products at a low temperature (<À70°C) in all these phenol O-glycosylation reactions, which are transformed into the final products at a higher temperature. BF 3 ÁOEt 2 is an effective catalyst to promote the conversion of 1,2-orthoesters into the corresponding 1,2-trans-O-glycosides. However, the 1,2-orthoesters could be converted into the dioxolenium triflate and glycosyl triflate in the presence of TMSOTf, these intermediates which might be in equilibrium with the glycosyl oxocarbenium related species lead to the final mixture of the a/b-O-glycosides and side-products.

Bioorganic & Medicinal Chemistry Letters, 2013

The biological role of installing a critical exocyclic enone into the structure of the alkaloid, ... more The biological role of installing a critical exocyclic enone into the structure of the alkaloid, (À)-eburnamonine, and characterization of the new chemical reactivity by quantitative NMR without using deuterated solvents are described. This selective modification to a natural product imparts potent anticancer activity as well as bestows chemical reactivity toward nucleophilic thiols, which was measured by quantitative NMR. The synthetic strategy provides an overall conversion of 40%. In the key synthetic step, a modified Peterson olefination was accomplished through the facile release of trifluoroacetate to create the requisite enone in the presence of substantial steric hindrance. j o u r n a l h o m e p a g e : w w w . e l s e v i e r . c o m / l o c a t e / b m c l activity in HL-60 (human leukemia) and cytotoxicity in MDA-MB-231 (breast cancer) cells (LC 50 = 14.1 lM); however, the parent