Publications by authors named "Chunhua Yuan"

59 Publications

Grincamycins P-T: Rearranged Angucyclines from the Marine Sediment-Derived sp. CNZ-748 Inhibit Cell Lines of the Rare Cancer Pseudomyxoma Peritonei.

J Nat Prod 2021 Apr 26. Epub 2021 Apr 26.

Department of Chemistry and Biochemistry, University of South Carolina, Columbia, South Carolina 29208, United States.

While marine natural products have been investigated for anticancer drug discovery, they are barely screened against rare cancers. Thus, in our effort to discover potential drug leads against the rare cancer pseudomyxoma peritonei (PMP), which currently lacks effective drug treatments, we screened extracts of marine actinomycete bacteria against the PMP cell line ABX023-1. This effort led to the isolation of nine rearranged angucyclines from sp. CNZ-748, including five new analogues, namely, grincamycins P-T (-). The chemical structures of these compounds were unambiguously established based on spectroscopic and chemical analyses. Particularly, grincamycin R () possesses an -containing α-l-methylthio-aculose residue, which was discovered in nature for the first time. All of the isolated compounds were evaluated against four PMP cell lines and some exhibited low micromolar inhibitory activities. To identify a candidate biosynthetic gene cluster (BGC) encoding the grincamycins, we sequenced the genome of the producing strain, sp. CNZ-748, and compared the BGCs detected with those linked to the production of angucyclines with different aglycon structures.
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http://dx.doi.org/10.1021/acs.jnatprod.1c00179DOI Listing
April 2021

Effects and mechanism of gating modifier spider toxins on the hERG channel.

Toxicon 2021 Jan 17;189:56-64. Epub 2020 Nov 17.

Key Laboratory of Mental Health of the Ministry of Education, Guangdong-Hong Kong-Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Guangdong Key Laboratory of Psychiatric Disorders, Department of Neurobiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, 510515, China. Electronic address:

Jingzhaotoxin-I, -III, -IV, -XIII, and -35 (JZTX-I, -III, -IV, -XIII, and -35), gating modifier toxins isolated from the venom of the Chinese tarantula Chilobrachys Jingzhao, were reported to act on cardiac sodium channels and Kv channels. JZTX-I and JZTX-XIII inhibited the hERG channel with the IC value of 626.9 nM and 612.6 nM, respectively. JZTX-III, -IV, and -35 share high sequence similarity with JZTX-I and JZTX-XIII, but they showed much lower affinity on the hERG channel compared with JZTX-I and JZTX-XIII. The inhibitory potency of the above five toxins on the hERG channel was not in accordance with their affinity on the Nav1.5 and Kv2.1 channels, indicating that the bioactive surfaces of the five toxins interacting with hERG, Nav1.5 and Kv2.1 are at least in part different. Structure-function analysis of the gating modifier toxins suggested that the functional bioactive surface binding to the hERG channel consists of a conserved hydrophobic patch, surrounding acidic residues (Glu10 in JZTX-XIII, Glu11 in JZTX-I), and basic residues which may be different from residues binding to the Kv2.1 channel.
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http://dx.doi.org/10.1016/j.toxicon.2020.11.008DOI Listing
January 2021

Rational design of cell-permeable cyclic peptides containing a d-Pro-l-Pro motif.

Bioorg Med Chem 2020 10 18;28(20):115711. Epub 2020 Aug 18.

Department of Chemistry and Biochemistry and Ohio State Biochemistry Program, The Ohio State University, 484 West 12(th) Avenue, Columbus, OH 43210, USA. Electronic address:

Cyclic peptides are capable of binding to challenging targets (e.g., proteins involved in protein-protein interactions) with high affinity and specificity, but generally cannot gain access to intracellular targets because of poor membrane permeability. In this work, we discovered a conformationally constrained cyclic cell-penetrating peptide (CPP) containing a d-Pro-l-Pro motif, cyclo(AFΦrpPRRFQ) (where Φ is l-naphthylalanine, r is d-arginine, and p is d-proline). The structural constraints provided by cyclization and the d-Pro-l-Pro motif permitted the rational design of cell-permeable cyclic peptides of large ring sizes (up to 16 amino acids). This strategy was applied to design a potent, cell-permeable, and biologically active cyclic peptidyl inhibitor, cyclo(YVNFΦrpPRR) (where Y is l-phosphotyrosine), against the Grb2 SH2 domain. Multidimensional NMR spectroscopic and circular dichroism analyses revealed that the cyclic CPP as well as the Grb2 SH2 inhibitor assume a predominantly random coil structure but have significant β-hairpin character surrounding the d-Pro-l-Pro motif. These results demonstrate cyclo(AFΦrpPRRFQ) as an effective CPP for endocyclic (insertion of cargo into the CPP ring) or exocyclic delivery of biological cargos (attachment of cargo to the Gln side chain).
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http://dx.doi.org/10.1016/j.bmc.2020.115711DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7577319PMC
October 2020

4,15-Dimethyl-7,12-diazo-niatri-cyclo-[10.4.0.0]hexa-deca-1(12),2,4,6,13,15-hexa-ene dibromide monohydrate.

Acta Crystallogr E Crystallogr Commun 2020 Sep 18;76(Pt 9):1467-1471. Epub 2020 Aug 18.

Department of Chemistry, University of Kentucky, 505 Rose Street, Lexington, Kentucky, 40506, USA.

The title compound, CHN·2Br·HO () is a member of the class of compounds called viologens. Viologens are quaternary salts of di-pyridyls and are especially useful as redox indicators as a result of their large negative one-electron reduction potentials. Compound consists of a dication composed of a pair of 4-methyl-pyridine rings mutually joined at the 2-position, with a dihedral angle between the pyridine rings of 62.35 (4)°. In addition, the rings are tethered the pyridine nitro-gen atoms by a tetra-methyl-ene bridge. Charge balance is provided by a pair of bromide anions, which are hydrogen bonded to a single water mol-ecule [ = 3.3670 (15) and 3.3856 (15) Å]. The crystal structure of , details of an improved synthesis, and a full analysis of its NMR spectra are presented.
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http://dx.doi.org/10.1107/S2056989020011147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7472763PMC
September 2020

Neuroprotection Against Parkinson's Disease Through the Activation of Akt/GSK3β Signaling Pathway by Tovophyllin A.

Front Neurosci 2020 9;14:723. Epub 2020 Jul 9.

Key Laboratory of Mental Health of the Ministry of Education, Guangdong-Hong Kong-Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Guangdong Province Key Laboratory of Psychiatric Disorders, Department of Neurobiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China.

Parkinson's disease (PD) is one of the most prevalent and life-threatening neurodegenerative disease and mainly characterized by lack of sufficient dopaminergic neurons in the substantia nigra pars compacta (SNc). Although current treatments help to alleviate clinical symptoms, effective therapies preventing neuronal loss remain scarce. Tovophyllin A (TA), one of the xanthones extracted from L. (GM), has recently been reported to play a beneficial role in the therapy of neurodegenerative diseases. In our research, we explored whether TA has protective effects on dopaminergic neurons in PD models. We found that TA significantly reduced apoptotic cell death in primary cortical neurons treated with 1-methyl-4-phenyl pyridinium (MPP) or paraquat (PQ) in the PD model. In an acute PD model induced by 1-methyl4-phenyl-1,2,3,5-tetrahydropyridine (MPTP) treatment, TA also attenuated the resulting behavioral dysfunctions and dopaminergic neuron loss. In the collected brain tissues, TA increased the phosphorylation of Akt and GSK-3β, which may be related to TA-mediated dopaminergic neuronal protective effects. In summary, our results illustrated that TA is a powerful cytoprotective agent for dopaminergic neurons in the MPTP-induced PD model, suggesting TA as a possible therapeutic candidate for PD.
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http://dx.doi.org/10.3389/fnins.2020.00723DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7364155PMC
July 2020

Cytotoxic and non-cytotoxic cardiac glycosides isolated from the combined flowers, leaves, and twigs of Streblus asper.

Bioorg Med Chem 2020 02 7;28(4):115301. Epub 2020 Jan 7.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, Columbus, OH 43210, United States. Electronic address:

A new non-cytotoxic [(+)-17β-hydroxystrebloside (1)] and two known cytotoxic [(+)-3'-de-O-methylkamaloside (2) and (+)-strebloside (3)] cardiac glycosides were isolated and identified from the combined flowers, leaves, and twigs of Streblus asper collected in Vietnam, with the absolute configuration of 1 established from analysis of its ECD and NMR spectroscopic data and confirmed by computational ECD calculations. A new 14,21-epoxycardanolide (3a) was synthesized from 3 that was treated with base. A preliminary structure-activity relationship study indicated that the C-14 hydroxy group and the C-17 lactone unit and the established conformation are important for the mediation of the cytotoxicity of 3. Molecular docking profiles showed that the cytotoxic 3 and its non-cytotoxic analogue 1 bind differentially to Na/K-ATPase. Compound 3 docks deeply in the Na/K-ATPase pocket with a sole pose, and its C-10 formyl and C-5, C-14, and C-4' hydroxy groups may form hydrogen bonds with the side-chains of Glu111, Glu117, Thr797, and Arg880 of Na/K-ATPase, respectively. However, 1 fits the cation binding sites with at least three different poses, which all depotentiate the binding between 1 and Na/K-ATPase. Thus, 3 was found to inhibit Na/K-ATPase, but 1 did not. In addition, the cytotoxic and Na/K-ATPase inhibitory 3 did not affect glucose uptake in human lung cancer cells, against which it showed potent activity, indicating that this cardiac glycoside mediates its cytotoxicity by targeting Na/K-ATPase but not by interacting with glucose transporters.
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http://dx.doi.org/10.1016/j.bmc.2019.115301DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7029422PMC
February 2020

Removal of hERG potassium channel affinity through introduction of an oxygen atom: Molecular insights from structure-activity relationships of strychnine and its analogs.

Toxicol Appl Pharmacol 2018 12 30;360:109-119. Epub 2018 Sep 30.

Key Laboratory of Mental Health of the Ministry of Education, Key Laboratory of Psychiatric Disorders of Guangdong Province, Department of Neurobiology, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China. Electronic address:

Nux vomica has been effectively used in Traditional Chinese Medicine. The processing of Nux vomica is necessary to reduce toxicity before it can be used in clinical practice. However, the mechanism for processing detoxification is unclear. hERG channels have been subjected to a routine test for compound cardiac toxicity in the drug development process. Therefore, we examined the effects and mechanisms of strychnine and brucine, two main ingredients of Nux vomica, and their N-oxides on hERG channels. Strychnine and brucine exhibited concentration-dependent inhibition of hERG channels with IC values of 25.9 μM and 44.18 μM, respectively. However, their nitrogen oxidative derivatives produced by processing of Nux vomica, strychnine N-oxide and brucine N-oxide, lost their activity on hERG channels. Compared to their parent compounds, only an oxygen atom was introduced in the nitrogen oxidative isoforms to compensate for the N - charge, suggesting that the protonated nitrogen is the key group for strychnine and brucine binding to hERG channel. Alanine-mutagenesis identified Y652 is the most important residue for strychnine and brucine binding to hERG channel. Y652A mutation increased the IC for strychnine and brucine by 21.64-fold and 29.78-fold that of WT I, respectively. Docking simulations suggested that the protonated nitrogen of strychnine and brucine formed a cation-π interaction with the aromatic ring of Y652. This study suggests that introduction of an oxygen to compensate for the N - charge could be a useful strategy for reducing hERG potency and increasing the safety margin of alkaloid-type compounds in drug development.
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http://dx.doi.org/10.1016/j.taap.2018.09.042DOI Listing
December 2018

Resonance assignments of wild-type and two cysteine-free variants of the four-helix bundle protein, Rop.

Biomol NMR Assign 2018 10 29;12(2):345-350. Epub 2018 Aug 29.

Department of Chemistry & Biochemistry, The Ohio State University, 100 W. 18th Ave., Columbus, OH, 43210, USA.

Repressor of primer (Rop, or ROM, RNA I modulator) is a 63 amino acid four-helix bundle protein that exists in solution as an anti-parallel homodimer. This protein has been extensively studied, including by X-ray crystallography, NMR, rational design, and combinatorial mutagenesis. Previous NMR experiments with wild-type Rop were carried out at pH 2.3 and pH 6.3. In this paper, we report complete N-H backbone assignments for three variants of Rop under the same pH 6.3 conditions: wild-type Rop; a cysteine-free pseudo-wild type variant (C38A C52V); and a core-repacked variant of the Cys-free variant (T19V L41V C38A C52V). These assignments enable functional and dynamic studies of wild-type and Cys-free variants of Rop.
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http://dx.doi.org/10.1007/s12104-018-9837-0DOI Listing
October 2018

Cytotoxic and NF-κB and mitochondrial transmembrane potential inhibitory pentacyclic triterpenoids from Syzygium corticosum and their semi-synthetic derivatives.

Bioorg Med Chem 2018 08 17;26(15):4452-4460. Epub 2018 Jul 17.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, Columbus, OH 43210, United States. Electronic address:

Syzygium is a large genus of flowering plants, with several species, including the clove tree, used as important resources in the food and pharmaceutical industries. In our continuing search for anticancer agents from higher plants, a chloroform extract of the leaves and twigs of Syzygium corticosum collected in Vietnam was found to be active toward the HT-29 human colon cancer cell line. Separation of this extract guided by HT-29 cells and nuclear factor-kappa B (NF-κB) inhibition yielded 19 known natural products, including seven triterpenoids, three ellagic acid derivatives, two methylated flavonoids, a cyclohexanone, four megastigmanes, a small lactone, and an aromatic aldehyde. The full stereochemistry of (+)-fouquierol (2) was defined for the first time. Biological investigations showed that (+)-ursolic acid (1) is the major cytotoxic component of S. corticosum, which exhibited also potent activities in the NF-κB and mitochondrial transmembrane potential (MTP) inhibition assays conducted, with IC values of 31 nM and 3.5 µM, respectively. Several analogues of (+)-ursolic acid (1) were synthesized, and a preliminary structure-activity relationship (SAR) study indicated that the C-3 hydroxy and C-28 carboxylic acid groups and 19,20-dimethyl substitution are all essential in the mediation of the bioactivities observed for this triterpenoid.
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http://dx.doi.org/10.1016/j.bmc.2018.07.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6177235PMC
August 2018

The Intracellular Loop of the Na/Ca Exchanger Contains an "Awareness Ribbon"-Shaped Two-Helix Bundle Domain.

Biochemistry 2018 08 3;57(34):5096-5104. Epub 2018 Jul 3.

Department of Chemistry and Biochemistry , The Ohio State University , Columbus , Ohio 43210 , United States.

The Na/Ca exchanger (NCX) is a ubiquitous single-chain membrane protein that plays a major role in regulating the intracellular Ca homeostasis by the counter transport of Na and Ca across the cell membrane. Other than its prokaryotic counterpart, which contains only the transmembrane domain and is self-sufficient as an active ion transporter, the eukaryotic NCX protein possesses in addition a large intracellular loop that senses intracellular calcium signals and controls the activation of ion transport across the membrane. This provides a necessary layer of regulation for the more complex function of eukaryotic cells. The Ca sensor in the intracellular loop is known as the Ca-binding domain (CBD12). However, how the signaling of the allosteric intracellular Ca binding propagates and results in transmembrane ion transportation still lacks a detailed explanation. Further structural and dynamics characterization of the intracellular loop flanking both sides of CBD12 is therefore imperative. Here, we report the identification and characterization of another structured domain that is N-terminal to CBD12 in the intracellular loop using solution nuclear magnetic resonance (NMR) spectroscopy. The atomistic structure of this domain reveals that two tandem long α-helices, connected by a short linker, form a stable crossover two-helix bundle (THB), resembling an "awareness ribbon". Considering the highly conserved amino acid sequence of the THB domain, the detailed structural and dynamics properties of the THB domain will be common among NCXs from different species and will contribute toward the understanding of the regulatory mechanism of eukaryotic Na/Ca exchangers.
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http://dx.doi.org/10.1021/acs.biochem.8b00300DOI Listing
August 2018

H, C, N resonance assignment of recombinant Euplotes raikovi protein Er-23.

Biomol NMR Assign 2018 10 6;12(2):291-295. Epub 2018 Jun 6.

Department of Chemistry & Biochemistry, The Ohio State University, 100 W. 18th Ave., Columbus, OH, 43210, USA.

Er-23 is a small, 51 amino acid, disulfide-rich pheromone protein used for cell signaling by Euplotes raikovi. Ten of the 51 amino acids are cysteine, allowing up to five disulfide bonds. Previous NMR work with Er-23 utilized homologously expressed protein, prohibiting isotopic labeling, and consequently the chemical shift assignments were incomplete. We have expressed uniformly N and C-labeled Er-23 in an E. coli expression system. Here we report the full backbone and side chain resonance assignments for recombinant Er-23.
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http://dx.doi.org/10.1007/s12104-018-9825-4DOI Listing
October 2018

Bioassay-Guided Isolation of Antioxidant and Cytoprotective Constituents from a Maqui Berry (Aristotelia chilensis) Dietary Supplement Ingredient As Markers for Qualitative and Quantitative Analysis.

J Agric Food Chem 2017 Oct 26;65(39):8634-8642. Epub 2017 Sep 26.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University , 500 West 12th Avenue, Columbus, Ohio 43210, United States.

Bioassay-guided phytochemical investigation of a commercially available maqui berry (Aristotelia chilensis) extract used in botanical dietary supplement products led to the isolation of 16 compounds, including one phenolic molecule, 1, discovered for the first time from a natural source, along with several known compounds, 2-16, including three substances not reported previously in A. chilensis, 2, 14, and 15. Each isolate was characterized by detailed analysis of NMR spectroscopic and HRESIMS data and tested for their in vitro hydroxyl radical scavenging and quinone-reductase inducing biological activities. A sensitive and accurate LC-DAD-MS method for the quantitative determination of the occurrence of six bioactive compounds, 6, 7, 10-12, and 14, was developed and validated using maqui berry isolates purified in the course of this study as authentic standards. The method presented can be utilized for dereplication efforts in future natural product research projects or to evaluate chemical markers for quality assurance and batch-to-batch standardization of this botanical dietary supplement component.
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http://dx.doi.org/10.1021/acs.jafc.7b03261DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5685509PMC
October 2017

Isolation of Bioactive Rotenoids and Isoflavonoids from the Fruits of Millettia caerulea.

Planta Med 2016 Jul 9;82(11-12):1096-104. Epub 2016 Jun 9.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, Columbus, Ohio, United States.

Three new rotenoids (1-3), two new isoflavonoids (4 and 5), and six known analogues (6-11) were isolated from an n-hexane partition of a methanol extract of the fruits of Millettia caerulea, with the structures of the new compounds elucidated by analysis of their spectroscopic data. The relative configurations of the rotenoids were determined by interpretation of their NMR spectroscopic data, and their absolute configurations were established using electronic circular dichroism spectra and specific rotation values. All compounds isolated were evaluated for their cell growth inhibitory activity against the HT-29 human colon cancer cell line, and the known compounds, (-)-3-hydroxyrotenone (6) and (-)-rotenone (7), were found to be potently active. When tested in an NF-κB inhibition assay, compound 6 showed activity. This compound, along with the new compound, (-)-caeruleanone D (1), and the known compound, ichthynone (8), exhibited K-Ras inhibitory potency. Further bioactivity studies showed that the new compounds, (-)-3-deoxycaeruleanone D (2) and (-)-3-hydroxycaeruleanone A (3), and the known compounds 8 and 11 induced quinone reductase in murine Hepa 1c1c7 cells.
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http://dx.doi.org/10.1055/s-0042-108059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4956498PMC
July 2016

Structural characterization of NRAS isoform 5.

Protein Sci 2016 May 24;25(5):1069-74. Epub 2016 Mar 24.

The Ohio State University Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio.

It was recently discovered that the NRAS isoform 5 (20 amino acids) is expressed in melanoma and results in a more aggressive cell phenotype. This novel isoform is responsible for increased phosphorylation of downstream targets such as AKT, MEK, and ERK as well as increased cellular proliferation. This structure report describes the NMR solution structure of NRAS isoform 5 to be used as a starting point to understand its biophysical interactions. The isoform is highly flexible in aqueous solution, but forms a helix-turn-coil structure in the presence of trifluoroethanol as determined by NMR and CD spectroscopy.
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http://dx.doi.org/10.1002/pro.2916DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4838646PMC
May 2016

Isolation and Structural Elucidation of Brevibacillin, an Antimicrobial Lipopeptide from Brevibacillus laterosporus That Combats Drug-Resistant Gram-Positive Bacteria.

Appl Environ Microbiol 2016 May 18;82(9):2763-2772. Epub 2016 Apr 18.

Department of Food Science and Technology, The Ohio State University, Columbus, Ohio, USA

A new environmental bacterial strain exhibited strong antimicrobial characteristics against methicillin-resistant Staphylococcus aureus, vancomycin-resistant strains of Enterococcus faecalis and Lactobacillus plantarum, and other Gram-positive bacteria. The producer strain, designated OSY-I1, was determined to be Brevibacillus laterosporusvia morphological, biochemical, and genetic analyses. The antimicrobial agent was extracted from cells of OSY-I1 with isopropanol, purified by high-performance liquid chromatography, and structurally analyzed using mass spectrometry (MS) and nuclear magnetic resonance (NMR). The MS and NMR results, taken together, uncovered a linear lipopeptide consisting of 13 amino acids and an N-terminal C6 fatty acid (FA) chain, 2-hydroxy-3-methylpentanoic acid. The lipopeptide (FA-Dhb-Leu-Orn-Ile-Ile-Val-Lys-Val-Val-Lys-Tyr-Leu-valinol, where Dhb is α,β-didehydrobutyric acid and valinol is 2-amino-3-methyl-1-butanol) has a molecular mass of 1,583.0794 Da and contains three modified amino acid residues: α,β-didehydrobutyric acid, ornithine, and valinol. The compound, designated brevibacillin, was determined to be a member of a cationic lipopeptide antibiotic family. In addition to its potency against drug-resistant bacteria, brevibacillin also exhibited low MICs (1 to 8 μg/ml) against selected foodborne pathogenic and spoilage bacteria, such as Listeria monocytogenes,Bacillus cereus, and Alicyclobacillus acidoterrestris Purified brevibacillin showed no sign of degradation when it was held at 80 °C for 60 min, and it retained at least 50% of its antimicrobial activity when it was held for 22 h under acidic or alkaline conditions. On the basis of these findings, brevibacillin is a potent antimicrobial lipopeptide which is potentially useful to combat drug-resistant bacterial pathogens and foodborne pathogenic and spoilage bacteria.
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http://dx.doi.org/10.1128/AEM.00315-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4836408PMC
May 2016

Structure of the Brd4 ET domain bound to a C-terminal motif from γ-retroviral integrases reveals a conserved mechanism of interaction.

Proc Natl Acad Sci U S A 2016 Feb 8;113(8):2086-91. Epub 2016 Feb 8.

Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210;

The bromodomain and extraterminal domain (BET) protein family are promising therapeutic targets for a range of diseases linked to transcriptional activation, cancer, viral latency, and viral integration. Tandem bromodomains selectively tether BET proteins to chromatin by engaging cognate acetylated histone marks, and the extraterminal (ET) domain is the focal point for recruiting a range of cellular and viral proteins. BET proteins guide γ-retroviral integration to transcription start sites and enhancers through bimodal interaction with chromatin and the γ-retroviral integrase (IN). We report the NMR-derived solution structure of the Brd4 ET domain bound to a conserved peptide sequence from the C terminus of murine leukemia virus (MLV) IN. The complex reveals a protein-protein interaction governed by the binding-coupled folding of disordered regions in both interacting partners to form a well-structured intermolecular three-stranded β sheet. In addition, we show that a peptide comprising the ET binding motif (EBM) of MLV IN can disrupt the cognate interaction of Brd4 with NSD3, and that substitutions of Brd4 ET residues essential for binding MLV IN also impair interaction of Brd4 with a number of cellular partners involved in transcriptional regulation and chromatin remodeling. This suggests that γ-retroviruses have evolved the EBM to mimic a cognate interaction motif to achieve effective integration in host chromatin. Collectively, our findings identify key structural features of the ET domain of Brd4 that allow for interactions with both cellular and viral proteins.
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http://dx.doi.org/10.1073/pnas.1516813113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4776507PMC
February 2016

Computer-Assisted Structure Elucidation of Black Chokeberry (Aronia melanocarpa) Fruit Juice Isolates with a New Fused Pentacyclic Flavonoid Skeleton.

Org Lett 2015 Jun 1;17(12):2988-91. Epub 2015 Jun 1.

†Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, 500 West 12th Avenue, Columbus, Ohio 43210, United States.

Melanodiol 4″-O-protocatechuate (1) and melanodiol (2) represent novel flavonoid derivatives isolated from a botanical dietary supplement ingredient, dried black chokeberry (Aronia melanocarpa) fruit juice. These noncrystalline compounds possess an unprecedented fused pentacyclic core with two contiguous hemiketals. Due to having significant hydrogen deficiency indices, their structures were determined using computer-assisted structure elucidation software. The in vitro hydroxyl radical-scavenging and quinone reductase-inducing activity of each compound are reported, and a plausible biogenetic scheme is proposed.
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http://dx.doi.org/10.1021/acs.orglett.5b01284DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4690212PMC
June 2015

pH dependence of the stress regulator DksA.

PLoS One 2015 23;10(3):e0120746. Epub 2015 Mar 23.

Department of Microbiology, The Ohio State University, Columbus, Ohio, United States of America; The Center for RNA Biology, The Ohio State University, Columbus, Ohio, United States of America.

DksA controls transcription of genes associated with diverse stress responses, such as amino acid and carbon starvation, oxidative stress, and iron starvation. DksA binds within the secondary channel of RNA polymerase, extending its long coiled-coil domain towards the active site. The cellular expression of DksA remains constant due to a negative feedback autoregulation, raising the question of whether DksA activity is directly modulated during stress. Here, we show that Escherichia coli DksA is essential for survival in acidic conditions and that, while its cellular levels do not change significantly, DksA activity and binding to RNA polymerase are increased at lower pH, with a concomitant decrease in its stability. NMR data reveal pH-dependent structural changes centered at the interface of the N and C-terminal regions of DksA. Consistently, we show that a partial deletion of the N-terminal region and substitutions of a histidine 39 residue at the domain interface abolish pH sensitivity in vitro. Together, these data suggest that DksA responds to changes in pH by shifting between alternate conformations, in which competing interactions between the N- and C-terminal regions modify the protein activity.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0120746PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4370453PMC
February 2016

Enhancement of Bacillus thuringiensis insecticidal activity by combining Cry1Ac and bi-functional toxin HWTX-XI from spider.

J Invertebr Pathol 2016 Mar 23;135:60-2. Epub 2015 Feb 23.

College of Life Science, Hunan Normal University, Changsha 410081, PR China. Electronic address:

In order to assess the potency of bi-functional HWTX-XI toxin from spider Ornithoctonus huwena in improving the insecticidal activity of Bacillus thuringiensis, a fusion gene of cry1Ac and hwtx-XI was constructed and expressed in an acrystalliferous B. thuringiensis strain Cry(-)B. Western blot analysis and microscopic observation revealed that the recombinant strain could express 140-kDa Cry1Ac-HWTX-XI fusion protein and produce parasporal inclusions during sporulation. Bioassay using the larvae of Helicoverpa armigera and Spodoptera exigua showed that the Cry1Ac-HWTX-XI fusion was more toxic than the control Cry1Ac protoxin, as revealed by 95% lethal concentration. Our study indicated that the HWTX-XI from spider might be a candidate for enhancing the toxicity of B. thuringiensis products.
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http://dx.doi.org/10.1016/j.jip.2015.02.005DOI Listing
March 2016

Cytotoxic and natural killer cell stimulatory constituents of Phyllanthus songboiensis.

Phytochemistry 2015 Mar 14;111:132-40. Epub 2015 Jan 14.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, Columbus, OH 43210, USA. Electronic address:

A dichapetalin-type triterpenoid and a dibenzylbutyrolactone-type lignan, together with five known lignans, a known aromatic diterpenoid, and a known acylated phytosterol, were isolated from the aerial parts of Phyllanthus songboiensis, collected in Vietnam. Their structures were determined by interpretation of the spectroscopic data, and the inhibitory activity toward HT-29 human colon cancer cells of all isolates was evaluated by a cytotoxicity assay. The known arylnaphthalene lignan, (+)-acutissimalignan A, was highly cytotoxic toward HT-29 cells, with an IC50 value of 19 nM, but this compound was inactive as a DNA topoisomerase IIα (topo IIα) poison. The known phytosterol, (-)-β-sitosterol-3-O-β-D-(6-O-palmitoyl)glucopyranoside, was found to stimulate natural killer (NK) cells at a concentration of 10μM in the presence of interleukin 12 (IL-12).
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http://dx.doi.org/10.1016/j.phytochem.2014.12.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4333069PMC
March 2015

Jingzhaotoxin-35, a novel gating-modifier toxin targeting both Nav1.5 and Kv2.1 channels.

Toxicon 2014 Dec 15;92:90-6. Epub 2014 Oct 15.

Key Laboratory of Psychiatric Disorders of Guangdong Province, Department of Neurobiology, Southern Medical University, Guangzhou 510515, China. Electronic address:

Jingzhaotoxin-35 (JZTX-35), a 36-residue polypeptide, was purified from the venom of the Chinese tarantula Chilobrachys jingzhao. JZTX-35 inhibited Nav1.5 and Kv2.1 currents with the IC50 value of 1.07 μM and 3.62 μM, respectively, but showed no significant effect on either Na(+) currents or Ca(2+) currents evoked in hippocampal neurons. It shifted the activation of the Nav1.5 and Kv2.1 channels to more depolarized voltages, and markedly shifted the steady-state inactivation of Nav1.5 currents toward more hyperpolarized potentials. Moreover, JZTX-35 can bind to a close state of Nav1.5 and Kv2.1 channels. These results indicate that JZTX-35 is a new gating modifier toxin. JZTX-35 shares high sequence similarity with Jingzhaotoxins (JZTXs) targeting Nav1.5 or Kv2.1 channels, but they showed different ion channel selectivity. Structure-function analysis in this study would provide important clues for the exploration of ion channel selectivity of JZTXs.
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http://dx.doi.org/10.1016/j.toxicon.2014.10.008DOI Listing
December 2014

Constituents of an extract of Cryptocarya rubra housed in a repository with cytotoxic and glucose transport inhibitory effects.

J Nat Prod 2014 Mar 17;77(3):550-6. Epub 2013 Dec 17.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University , Columbus, Ohio 43210, United States.

A new alkylated chalcone (1), a new 1,16-hexadecanediol diester (2), and eight known compounds were isolated from a dichloromethane-soluble repository extract of the leaves and twigs of Cryptocarya rubra collected in Hawaii. The structures of the new compounds were determined by interpretation of their spectroscopic data, and the absolute configurations of the two known cryptocaryanone-type flavonoid dimers, (+)-bicaryanone A (3) and (+)-chalcocaryanone C (4), were ascertained by analysis of their electronic circular dichroism and NOESY NMR spectra. All compounds isolated were evaluated against HT-29 human colon cancer cells, and, of these, (+)-cryptocaryone (5) was found to be potently cytotoxic toward this cancer cell line, with an IC50 value of 0.32 μM. This compound also exhibited glucose transport inhibitory activity when tested in a glucose uptake assay.
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http://dx.doi.org/10.1021/np400809wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4047178PMC
March 2014

Sphenostylisins A-K: bioactive modified isoflavonoid constituents of the root bark of Sphenostylis marginata ssp. erecta.

J Org Chem 2013 Oct 3;78(20):10166-77. Epub 2013 Oct 3.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University , 500 West 12th Avenue, Columbus, Ohio 43210, United States.

Sphenostylisins A-C (1-3), three complex dimeric compounds representing two novel carbon skeletons, along with an additional eight new compounds, sphenostylisins D-K (4-11), were isolated from the active chloroform-soluble extract of the root bark of Sphenostylis marginata ssp. erecta using a bioactivity-guided isolation approach. The structures were elucidated by means of detailed spectroscopic analysis, including NMR and HRESIMS analysis, and tandem MS fragmentation was utilized to further support the structures of 1-3. The absolute configuration of sphenostylisin C (3) was established by electronic circular dichroism analysis. Plausible biogenetic relationships between the modified isoflavonoids 1-11 are proposed, and a cyclization reaction of 9 was conducted to support one of the biogenetic proposals made. All of these pure isolates were evaluated against a panel of in vitro bioassays, and among the results obtained, sphenostylisin A (1) was found to be a very potent NF-κB inhibitor (IC50 = 6 nM).
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http://dx.doi.org/10.1021/jo401573hDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3827909PMC
October 2013

The study of pH-dependent stability shows that the TPLH-mediated hydrogen-bonding network is important for the conformation and stability of human gankyrin.

Biochemistry 2013 Jul 8;52(28):4848-57. Epub 2013 Jul 8.

Campus Chemical Instrument Center, §Ohio State Biochemistry Program, ∥Division of Environmental Health Sciences, College of Public Health, and ‡Comprehensive Cancer Center, The Ohio State University , Columbus, Ohio 43210, United States.

Ankyrin repeat (AR) proteins possess a distinctive modular and repetitive architecture, and their global folds are maintained by short-range interactions in terms of the primary sequence. In this work, we extended our previous study on the highly conserved TPLH tetrapeptide and investigated the impact of a solvent-exposed histidine residue on the pH-dependent stability of gankyrin, providing further insight into the contribution of the TPLH motif to the tertiary fold of AR proteins. Consisting of seven ARs, gankyrin has five histidine residues in TPLH motifs or its variants, all of which adopt a H(ε2)-tautermeric form and are shielded from solvent. By truncating the C-terminal ankyrin repeat (AR7), we exposed H177 in the (174)TPLH(177) of AR6 (the second C-terminal AR) to an aqueous environment. We showed that this truncated gankyrin mutant, namely, Gank(1-201), was well-folded at a neutral pH with a slightly lower stability with respect to gankyrin wild type (WT). However, unlike gankyrin WT, the stability of Gank(1-201) was markedly decreased together with a loss of conformation at a pH slightly below 6.0. It was rationalized that the protonation of the H177 imidazole ring triggered the disruption of the TPLH-mediated hydrogen-bonding network, which in turn led to the loss of conformation and stability. These results together with the work on Q210H mutant nicely explain that the C-terminal AR7 has a (207)TPLQ(210) variant and are in support of the notion that a string of TPLH/variant, which may arguably act like a zip lock to the elongated AR proteins via intra-/inter-repeat hydrogen-bonding, is important in maintaining the tertiary fold. Additionally, we made rational mutagenesis to introduce extra surface charge on AR7 of gankyrin and demonstrated that G214E and I219D mutations increased the stability of gankyrin while the function remained intact. Taken together, our results indicate that the TPLH-mediated hydrogen-bonding network is important for the conformation and stability of human gankyrin, and the C-terminal AR contributes to the conformational stability of gankyrin (AR proteins in general) through shielding this TPLH network from solvent as well as making the surface area more accessible to solvent.
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http://dx.doi.org/10.1021/bi4005717DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3843994PMC
July 2013

Evidence that P12, a specific variant of P16(INK4A), plays a suppressive role in human pancreatic carcinogenesis.

Biochem Biophys Res Commun 2013 Jun 29;436(2):217-22. Epub 2013 May 29.

Department of Pharmacy, The Arthur G. James Cancer Hospital and Richard J. Solove Research Institute, The Ohio State University, Columbus, OH 43210, United States.

The INK4a-ARF locus plays a central role in the development of pancreatic tumors as evidenced by the fact that up to 98% of pancreatic tumor specimens harbored genetic alterations at the INK4a-ARF locus. Interestingly, in addition to the well-known P16(INK4A) (P16) and P14ARF tumor suppressors, the INK4a-ARF locus in pancreas encodes another protein, P12, whose structure, function, and contributions to pancreatic carcinogenesis remain to be elucidated. In the current study, we demonstrated that over-expression of p12 in human pancreatic cancer cells led to cell arrest at the G1 phase and such cell cycle arrest was related to down-regulation of a number of oncogenes, such as c-Jun, Fos, and SEI1. Furthermore, unlike P16, P12 did not retain any cyclin-dependent kinase 4 (CDK4)-inhibitory activity. Instead, P12 exhibited a transactivating activity not found in P16. We also examined the genetic status of p12 in a cohort of 40 pancreatic tumor specimens and found that p12 alteration was prevalent in pancreatic tumors with an incidence of 70% (28/40). These results support that P12 is a tumor suppressive protein distinct from P16, and its genetic inactivation is associated with pancreatic carcinogenesis.
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http://dx.doi.org/10.1016/j.bbrc.2013.05.078DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3752424PMC
June 2013

An Arabidopsis cell wall proteoglycan consists of pectin and arabinoxylan covalently linked to an arabinogalactan protein.

Plant Cell 2013 Jan 31;25(1):270-87. Epub 2013 Jan 31.

Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia 30602-4712, USA.

Plant cell walls are comprised largely of the polysaccharides cellulose, hemicellulose, and pectin, along with ∼10% protein and up to 40% lignin. These wall polymers interact covalently and noncovalently to form the functional cell wall. Characterized cross-links in the wall include covalent linkages between wall glycoprotein extensins between rhamnogalacturonan II monomer domains and between polysaccharides and lignin phenolic residues. Here, we show that two isoforms of a purified Arabidopsis thaliana arabinogalactan protein (AGP) encoded by hydroxyproline-rich glycoprotein family protein gene At3g45230 are covalently attached to wall matrix hemicellulosic and pectic polysaccharides, with rhamnogalacturonan I (RG I)/homogalacturonan linked to the rhamnosyl residue in the arabinogalactan (AG) of the AGP and with arabinoxylan attached to either a rhamnosyl residue in the RG I domain or directly to an arabinosyl residue in the AG glycan domain. The existence of this wall structure, named ARABINOXYLAN PECTIN ARABINOGALACTAN PROTEIN1 (APAP1), is contrary to prevailing cell wall models that depict separate protein, pectin, and hemicellulose polysaccharide networks. The modified sugar composition and increased extractability of pectin and xylan immunoreactive epitopes in apap1 mutant aerial biomass support a role for the APAP1 proteoglycan in plant wall architecture and function.
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http://dx.doi.org/10.1105/tpc.112.107334DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3584541PMC
January 2013

Antioxidant and quinone reductase-inducing constituents of black chokeberry (Aronia melanocarpa) fruits.

J Agric Food Chem 2012 Nov 13;60(46):11551-9. Epub 2012 Nov 13.

Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, 500 West 12th Avenue, Columbus, Ohio 43210, USA.

Using in vitro hydroxyl radical-scavenging and quinone reductase-inducing assays, bioactivity-guided fractionation of an ethyl acetate-soluble extract of the fruits of the botanical dietary supplement, black chokeberry (Aronia melanocarpa), led to the isolation of 27 compounds, including a new depside, ethyl 2-[(3,4-dihydroxybenzoyloxy)-4,6-dihydroxyphenyl] acetate (1), along with 26 known compounds (2-27). The structures of the isolated compounds were identified by analysis of their physical and spectroscopic data ([α](D), NMR, IR, UV, and MS). Altogether, 17 compounds (1-4, 9, 15-17, and 19-27) showed significant antioxidant activity in the hydroxyl radical-scavenging assay, with hyperin (24, ED(50) = 0.17 μM) being the most potent. The new compound (1, ED(50) = 0.44 μM) also exhibited potent antioxidant activity in this assay. Three constituents of black chokeberry fruits doubled quinone reductase activity at concentrations <20 μM, namely, protocatechuic acid [9, concentration required to double quinone reductase activity (CD) = 4.3 μM], neochlorogenic acid methyl ester (22, CD = 6.7 μM), and quercetin (23, CD = 3.1 μM).
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http://dx.doi.org/10.1021/jf303712eDOI Listing
November 2012

Isolation of a Paenibacillus sp. strain and structural elucidation of its broad-spectrum lipopeptide antibiotic.

Appl Environ Microbiol 2012 May 24;78(9):3156-65. Epub 2012 Feb 24.

Department of Food Science & Technology, Ohio State University, Columbus, Ohio, USA.

This research was initiated to search for novel antimicrobial compounds produced by food or environmental microorganisms. A new bacterial strain, designated OSY-SE, which produces a unique and potent antimicrobial agent was isolated from soil. The isolate was identified as a Paenibacillus sp. through cultural, biochemical, and genetic analyses. An antimicrobial compound was extracted from Paenibacillus OSY-SE with acetonitrile and purified using liquid chromatography. After analyses by mass spectrometry (MS) and nuclear magnetic resonance (NMR), the antimicrobial compound was determined to be a cyclic lipopeptide consisting of a C(15) fatty acyl (FA) chain and 13 amino acids. The deduced sequence is FA-Orn-Val-Thr-Orn-Ser-Val-Lys-Ser-Ile-Pro-Val-Lys-Ile. The carboxyl-terminal Ile is connected to Thr by ester linkage. The new compound, designated paenibacterin, showed antagonistic activities against most Gram-positive and Gram-negative bacteria tested, including Listeria monocytogenes, methicillin-resistant Staphylococcus aureus, Escherichia coli O157:H7, and Salmonella enterica serovar Typhimurium. Paenibacterin is resistant to trypsin, lipase, α-glucosidase, and lysozyme. Its antimicrobial activity was lost after digestion by pronase and polymyxin acylase. Paenibacterin is readily soluble in water and fairly stable to exposure to heat and a wide range of pH values. The new isolate and its antimicrobial agent are being investigated for usefulness in food and medical applications.
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http://dx.doi.org/10.1128/AEM.07782-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3346447PMC
May 2012

JZTX-XIII, a Kv channel gating modifier toxin from Chinese tarantula Chilobrachys jingzhao.

Toxicon 2012 Feb 8;59(2):265-71. Epub 2011 Dec 8.

Department of Neurobiology, Southern Medical University, Guangzhou, PR China.

Jingzhaotoxin-XIII (JZTX-XIII), a 35 residue polypeptide, with the ability to inhibit voltage-dependent potassium channels in the shab (Kv2) and shal (Kv4) subfamilies, was purified from the venom of the Chinese tarantula Chilobrachys jingzhao. Electrophysiological recordings carried out in Xenopus laevis oocytes showed that JZTX-XIII acted as gating modifier of voltage-dependent K+ channels which inhibited the Kv2.1 channel and Kv4.1 channel, with the IC50 value of 0.47 μM and 1.17 μM, respectively. JZTX-XIII shares high sequence similarity with gating modifier toxins inhibiting a wide variety of ion channels including Nav1.5 subtype, but it showed no Nav1.5 channel activity. Structure-function analysis indicates that the acidic residues of Glu10 and Glu17 in JZTX-XIII might be responsible for the loss of the Nav1.5 channel inhibitory potency for JZTX-XIII.
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http://dx.doi.org/10.1016/j.toxicon.2011.11.021DOI Listing
February 2012

Simultaneous binding of two peptidyl ligands by a SRC homology 2 domain.

Biochemistry 2011 Sep 9;50(35):7637-46. Epub 2011 Aug 9.

Ohio State Biochemistry Program, The Ohio State University, Columbus, Ohio 43210, United States.

Src homology 2 (SH2) domains mediate protein-protein interactions by recognizing phosphotyrosine (pY)-containing sequences of target proteins. In all of the SH2 domain-pY peptide interactions described to date, the SH2 domain binds to a single pY peptide. Here, determination of the cocrystal structure of the N-terminal SH2 domain of phosphatase SHP-2 bound to a class IV peptide (VIpYFVP) revealed a noncanonical 1:2 (protein-peptide) complex. The first peptide binds in a canonical manner with its pY side chain inserted in the usual binding pocket, while the second pairs up with the first to form two antiparallel β-strands that extend the central β-sheet of the SH2 domain. This unprecedented binding mode was confirmed in the solution phase by NMR experiments and shown to be adopted by pY peptides derived from cellular proteins. Site-directed mutagenesis and surface plasmon resonance studies revealed that the binding of the first peptide is pY-dependent, but phosphorylation is not required for the second peptide. Our findings suggest a potential new function for the SH2 domain as a molecular clamp to promote dimerization of signaling proteins.
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http://dx.doi.org/10.1021/bi200439vDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3164524PMC
September 2011