1.
Identification of five gelatins by ultra performance liquid chromatography/time-of-flight mass spectrometry (UPLC/Q-TOF-MS) using principal component analysis.
Source
Research and Inspection Center of Traditional Chinese Medicine and Ethnomedicine, National Institutes for Food and Drug Control, State Food and Drug Administration, 2 Tiantan Xili, Beijing 100050, China; School of Chinese Pharmacy, Beijing University of Chinese Medicine, No. 6 Wangjing Zhong Huan Nan Lu, Chaoyang District, Beijing 100102, China.
Abstract
An ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry (UPLC/Q-TOF-MS) method coupled with a principal component analysis (PCA) was developed and applied toward identifying donkey-hide gelatin, bovine-hide gelatin, pig-hide gelatin, tortoise shell glue, and deerhorn glue. The UPLC-MS data of the trypsin digested samples were subjected to principal component analysis (PCA) in order to classify these five gelatins. Additionally, marker peptides given by the loadings plot of PCA were identified based on a comparison of recorded LC-MS data with a previously reported database of the corresponding gelatin variants. The results from this study indicate that the proposed method is reliable, and it has been successfully applied to the identification of variants of gelatins commonly used in Traditional Chinese Medicine (TCM).
Copyright © 2011 Elsevier B.V. All rights reserved.
In-silico characterization of ECE-1 inhibitors.
Source
R.G. Biosciences, Visakhapatnam, India.
Abstract
Atherosclerosis is the primary cause of CAD and cerebrovascular disease. Endothelin (ET)-1 is a vasoconstrictivepeptide implicated in Atherosclerosis pathology. Endothelin-converting enzyme (ECE) is a membrane metalloprotease that generates endothelin. Reported inhibitors of ECE-1 and their IC(50) values were retrieved from literature and their structures were docked with the parent protein using the Molegro virtual docker. The obtained MolDock scores of each of the compounds are hereby reported and are subject to graphical analysis in conjunction with their respective IC(50) values to characterize potent inhibitors. A search was then run in the ZINC database for compounds with similar properties. Potent inhibitors with higher Dock scores and better Ranking were isolated and are reported.
Copyright © 2012 Elsevier Ltd. All rights reserved.
Identification and classification of conopeptides using profile Hidden Markov Models.
Source
Estonian Biocentre, Riia 23, 51010, Tartu, Estonia.
Abstract
Conopeptides are small toxins produced by predatory marine snails of the genus Conus. They are studied with increasing intensity due to their potential in neurosciences and pharmacology. The number of existing conopeptides is estimated to be 1 million, but only about 1000 have been described to date. Thanks to new high-throughput sequencing technologies the number of known conopeptides is likely to increase exponentially in the near future. There is therefore a need for a fast and accurate computational method for identification and classification of the novel conopeptides in large data sets. 62 profile Hidden Markov Models (pHMMs) were built for prediction and classification of all described conopeptide superfamilies and families, based on the different parts of the corresponding protein sequences. These models showed very high specificity in detection of new peptides. 56 out of 62 models do not give a single false positive in a test with the entire UniProtKB/Swiss-Prot protein sequence database. Our study demonstrates the usefulness of mature peptide models for automatic classification with accuracy of 96% for the mature peptide models and 100% for the pro- and signal peptide models. Our conopeptide profile HMMs can be used for finding and annotation of new conopeptides from large datasets generated by transcriptome or genome sequencing. To our knowledge this is the first time this kind of computational method has been applied to predict all known conopeptide superfamilies and some conopeptide families.
Copyright © 2011. Published by Elsevier B.V.
Semi-Automated Identification of N-Glycopeptides by Hydrophilic Interaction Chromatography, nano-Reverse-Phase LC-MS/MS, and Glycan DatabaseSearch.
Abstract
Glycoproteins fulfill many indispensable biological functions and changes in protein glycosylation have been observed in various diseases. Improved analytical methods are needed to allow a complete characterization of this complex and common posttranslational modification. In this study, we present a workflow for the analysis of the microheterogeneity of N-glycoproteins which couples hydrophilic interaction and nano-reverse-phase C18 chromatography to tandem QTOF mass spectrometric analysis. A glycan database search program, GlycoPeptideSearch, was developed to match N-glycopeptide MS/MS spectra with the glycopeptides comprised of a glycan drawn from the GlycomeDB glycan structuredatabase and a peptide from a user-specified set of potentially glycosylated peptides. Application of the workflow to human haptoglobin and hemopexin, two microheterogeneous N-glycoproteins, identified a total of 57 distinct site-specific glycoforms in the case of haptoglobin and 14 site-specific glycoforms of hemopexin. Using glycan oxonium ions, peptide-characteristic glycopeptide fragment ions, and by collapsing topologically redundant glycans, the search software was able to make unique N-glycopeptide assignments for 51% of assigned spectra, with the remaining assignments primarily representing isobaric topological rearrangements. The optimized workflow, coupled with GlycoPeptideSearch, is expected to make high-throughput semi-automated glycopeptide identification feasible for a wide range of users.
Cy5.5-Gly-Pro-Leu-Gly-Val-Arg-Gly-(TDOPA)3-flower-like gold–Fe3O4 optical nanoparticles.
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Oct 06 [updated 2012 Jan 05].
Excerpt
Extracellular matrix (ECM) adhesion molecules consist of a complex network of fibronectins, collagens, chondroitins, laminins, glycoproteins, heparin sulfate, tenascins, and proteoglycans that surround connective tissue cells, and they are mainly secreted by fibroblasts, chondroblasts, and osteoblasts (1). Cell substrate adhesion molecules are considered essential regulators of cell migration, differentiation, and tissue integrity and remodeling. These molecules play a role in inflammation and atherogenesis, but they also participate in the process of invasion and metastasis of malignant cells in the host tissue (2). Tumor cells adhere to the ECM, which provides a matrix environment for permeation of tumor cells through the basal lamina and underlying interstitial stroma of the connective tissue. Overexpression of matrix metalloproteinases (MMPs) and other proteases by tumor cells allows intravasation of tumor cells into the circulatory system after degrading the basement membrane and ECM (3). Gold has not been used as an X-ray contrast agent in vivo. Gold has a higher atomic number and a higher absorption coefficient than iodine, providing 2.7-fold greater contrast/weight than iodine (4). Furthermore, imaging gold at 80–100 keV reduces interference from bone absorption and provides lower soft tissue absorption, which would reduce radiation to patients. Hainfeld et al. (4) used gold nanoparticles (AuNPs; 1.9 nm in diameter, ~50 kDa) as a computed tomography (CT) contrast agent in mice; these experiments showed enhanced CT contrast of the vasculature, kidneys, and tumors in mice. However, plasma proteins in blood adsorb onto the surface of bare AuNPs, which produces large aggregates (5) that may result in altered pharmacokinetics and biodistribution of AuNPs (6). Polyethylene glycol (PEG) is found to minimize nonspecific adsorption of proteins onto NPs and to reduce their uptake by the liver (6). PEG-AuNPs have been being studied as cancer CT imaging and photothermal agents (7). Several families of MMPs are involved in atherogenesis, myocardial infarction, angiogenesis, and tumor invasion and metastases (8-11). MMP expression is highly regulated in normal cells, such as trophoblasts, osteoclasts, neutrophils, and macrophages. Elevated levels of MMPs have been found in tumors associated with a poor prognosis for cancer patients (12). The peptide Gly-Pro-Leu-Gly-Val-Arg-Gly-Cys-NH2 was found to be a MMP substrate and is cleaved between the Leu and Gly residues. Lee et al. (13) used this sequence with a Cy5.5 NIR dye molecule to attach to AuNPs to form fluorescence-quenched nanoparticles (Cy5.5-Gly-Pro-Leu-Gly-Val-Arg-Gly-Cys-AuNPs (Cy5.5-MMP-AuNPs or GANPs). The Cy5.5 molecules are in close proximity, resulting in fluorescence quenching because of efficient fluorescence resonance energy transfer to Au. NIR fluorescence signal will increase when the Leu-Gly bond is cleaved by MMPs, releasing Cy5.5-containing fragments. Cy5.5 is a NIR fluorescent dye with an absorbance maximum at 675 nm, an emission maximum at 694 nm, and a high extinction coefficient of 250,000 M−1cm−1. Cy5.5-MMP-AuNPs are being developed for NIR fluorescence imaging of MMPs expressed in tumors, atherosclerosis, myocardial infarction, and other diseases. Xie et al. (14) replaced the AuNP with Au-Fe3O4, a composite NP shaped like a flower, to induce a fluorescently quenched state to the overall nanostructure. There were three iron oxide “petals” on each AuNP. The MMP peptide was covalently linked to an anchoring unit, Lys-tridihydrophenylalanine (Lys-TDOPA) on the surface of the iron oxide NP. The gold surface was passivated with a thiolated PEG (SH-PEG5000). Cy5.5-Gly-Pro-Leu-Gly-Val-Arg-Gly-(Lys-TDOPA)3-flower-like gold–Fe3O4 optical NPs (FANPs) have been evaluated for imaging protease expression in vivo.
Sections
18F-Labeled N-(4-fluorobenzylidene)oxime-dimeric (ZHER2:477)2 .
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Nov 02 [updated 2012 Jan 04].
Excerpt
The 18F-labeled N-(4-fluorobenzylidene)oxime (FBO)-dimeric (ZHER2:477)2 conjugate, abbreviated as 18F-FBO-(ZHER2:477)2, is an affibody derivative synthesized by Cheng et al. for positron emission tomography (PET) of HER2-expressing tumors (1). Affibody molecules are a group of nonimmunogenic scaffold proteins that derive from the B-domain of staphylococcal surface protein A (2, 3). In the past several years, affibodies have drawn significant attention for developing imaging and therapeutic agents because of their unique features (3, 4). First, affibodies are small, with only 58 amino acid residues (~7 kDa) (3, 5). The small size allows affibodies to be generated with solid-phase peptidesynthesis and to be cleared quickly from kidneys. Second, affibodies have a high binding affinity and specificity to their targets. Their binding affinity can be further improved by generating multimeric constructs through the solvent-exposed termini of affibody Z-domain. The anti-HER2 monomeric affibody ZHER2:4 is an example that has a binding affinity of ~50 nM, but its dimeric form, (ZHER2:4)2, exhibits an improved binding affinity of up to ~3 nM in vitro (6). Third, affibodies lack cysteine residues and disulfide bridges in structure, and they fold rapidly. These features make it possible to chemically synthesize fully functional molecules and to introduce unique cysteine residues or chemical groups into affibodies for site-specific labeling. Several anti-HER2 affibody derivatives have been synthesized in this way. The imaging agent HPEM-His6-(ZHER2:4)2-Cys was generated by radiobrominating the dimeric (ZHER2:4)2 through the cysteine residues that were introduced to the C-terminus of (ZHER2:4)2 (7). Several affibody derivatives (e.g., 68Ga-DOTA-ZHER2:342-pep2, 111In-DOTA-ZHER2:342-pep2, 111In-benzyl-DOTA-ZHER2:342, and 111In-benzyl-DTPA-ZHER2:342) were synthesized by coupling a chelating agent with a specifically protected site group of the ZHER2:342peptide chain (3). Furthermore, affibody proteins can be selected and optimized with a strategy of sequence mutation and affinity maturation, and an example selected with this strategy is the anti-HER2 affibody ZHER2:342, which has an increased affinity of 50 nM (ZHER2:4, the first generation) to 22 pM (8). The investigators at Stanford University first tested the feasibility of the monomeric and dimeric forms of anti-HER2 affibody ZHER2:477 for molecular imaging. Both forms of the ZHER2:477 molecule were radiofluorinated with an 18F-labeled prosthetic group of 4-18F-fluorobenzaldehyde (18F-FBO-ZHER2:477 and 18F-FBO-(ZHER2:477)2, respectively) (1). The investigators have also coupled 64Cu to the affibody through DOTA, leading to the development of imaging agents 64Cu-DOTA- ZHER2:477 and 64Cu-DOTA-(ZHER2:477)2 (9). Interestingly, these studies showed that smaller affibody constructs performed better in vivo in terms of tumor uptake and clearance in spite of the lower affinity in vitro. The investigators then generated a class of small proteins consisting of two α-helix bundles of the 3-helix affibody by deleting the helix 3 because the binding domain localizes in the α-helices 1 and 2 bundles (5). One of these 2-helix proteins is MUT-DS, which has α-helices 1 and 2 bundles, with a disulfide bridge being formed between the two inserted homocysteines (10-12). MUT-DS showed a binding affinity to HER2 in the low-nM range. The radiolabeled MUT-DS derivatives exhibited favorable pharmacokinetics for both imaging and therapy of HER2-expressing tumors (refer to MUT-DS derived agents in MICAD). This series of chapters summarizes the data obtained with the ZHER2:477 derivatives, and this chapter presents the data obtained with 18F-FBO-(ZHER2:477)2 (1).
Sections
VivoTag-S 750-(S)-2-amino-4-pentynoic acid12-exendin-4.
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Nov 01 [updated 2012 Jan 05].
Excerpt
Glucagon-like peptide-1 (GLP-1; 30 amino acids) is secreted from enteroendocrine cells of the distal small intestine in response to food ingestion (1). GLP-1 plays an important role in glucose metabolism and homeostasis by inhibiting gastric emptying, glucagon secretion, and glucose production (2). In addition, GLP-1 induces insulin release from the pancreatic β-cells as well as their proliferation. The GLP-1 receptor (GLP-1R) has been identified in normal tissues such as the pancreatic β-cells, stomach, brain, and lung, and it has been shown to be highly overexpressed in human insulinomas and gastrinomas (3). In insulinomas, GLP-1R density is considerably greater, and the GLP-1R is more frequently observed than somatostatin receptors. Exendin-4 is a GLP-1 analog with 39 amino acids isolated from the venom of the Gila monster (Heloderma suspectum) (4). Exendin-4 and GLP-1 share a 53% amino acid sequence homology. Exendin-4 is a more potent and longer-lasting GLP-1R agonist than GLP-1. Exendin-4 is resistant to cleavage by dipeptidyl peptidase IV, whereas the first two N-terminal amino acids of GLP-1 are rapidly cleaved. Exenatide, a synthetic version of exendin-4, is the first GLP-1 mimetic approved by the United States Food and Drug Administration (FDA) for use in select patients with type 2 diabetes (5). 111In-Labeled diethylenetriamine pentaacetic acid-aminohexanoic acid-Lys40-exendin-4 (111In-DTPA-Ahx-Lys40-exendin-4) has been developed for single-photon emission computed tomography (SPECT) imaging of the GLP-1R (6). For optical near-infrared (NIR) fluorescence imaging, Reiner et al. (7) replaced the Lys at position 12 of exendin-4 with the non-natural amino acid (S)-2-amino-4-pentynoic acid to form a neopeptide, E4X12. The azide-functionalized NIR dye VivoTag-S 750 (VT750) was conjugated to the non-natural amino acid at position 12 via azide-alkyne cycoaddition to form VT750-(S)-2-amino-4-pentynoic acid12-exendin-4 (E4X12-VT750). E4X12-VT750 has been evaluated as an optical imaging agent for GLP-1R density and β-cell mass (BCM) in normal mice and in mice treated with streptozotocin (STZ) to induce BCM loss and diabetes.
Sections
18F-Labeled N-(4-fluorobenzylidene)oxime-monomeric ZHER2:477 .
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Nov 02 [updated 2012 Jan 04].
Excerpt
The 18F-labeled N-(4-fluorobenzylidene)oxime (FBO)-monomeric ZHER2:477 conjugate, abbreviated as 18F-FBO-ZHER2:477, is an affibody derivative synthesized by Cheng et al. for positron emission tomography (PET) of HER2-expressing tumors (1). Affibody molecules are a group of nonimmunogenic scaffold proteins that derive from the B-domain of staphylococcal surface protein A (2, 3). In the past several years, affibodies have drawn significant attention for developing imaging and therapeutic agents because of their unique features (3, 4). First, affibodies are small, with only 58 amino acid residues (~7 kDa) (3, 5). The small size allows affibodies to be generated with solid-phase peptidesynthesis and to be cleared quickly by the kidneys. Second, affibodies have a high binding affinity and specificity to their targets. Their binding affinity can be further improved by generating multimeric constructs through the solvent-exposed termini of affibody Z-domain. The anti-HER2 monomeric affibody ZHER2:4 is an example that has a binding affinity of ~50 nM, but its dimeric form, (ZHER2:4)2, exhibits an improved binding affinity of up to ~3 nM (6). Third, affibodies lack cysteine residues and disulfide bridges in structure, and they fold rapidly. These features make it possible to chemically synthesize fully functional molecules and to introduce unique cysteine residues or chemical groups into affibodies for site-specific labeling. Several anti-HER2 affibody derivatives have been synthesized in this way. The imaging agent HPEM-His6-(ZHER2:4)2-Cys was generated by radiobrominating the dimeric (ZHER2:4)2 through the cysteine residues that were introduced to the C-terminus of (ZHER2:4)2 (7). Several affibody derivatives (e.g., 68Ga-DOTA-ZHER2:342-pep2, 111In-DOTA-ZHER2:342-pep2, 111In-benzyl-DOTA-ZHER2:342, and 111In-benzyl-DTPA-ZHER2:342) were synthesized by coupling a chelating agent with a specifically protected site group of the ZHER2:342peptide chain (3). Furthermore, these small affibody proteins can be selected and optimized with a strategy of sequence mutation and affinity maturation, and an example selected with this strategy is the anti-HER2 affibody ZHER2:342, which has an increased affinity of 50 nM (ZHER2:4, the first generation) to 22 pM (8). The investigators at Stanford University first tested the feasibility of the monomeric and dimeric forms of anti-HER2 affibody ZHER2:477 for molecular imaging. Both forms of the ZHER2:477 molecule were radiofluorinated with an 18F-labeled prosthetic group of 4-18F-fluorobenzaldehyde (18F-FBO-ZHER2:477 and 18F-FBO-(ZHER2:477)2, respectively) (1). The investigators have also coupled 64Cu to the affibody through DOTA, leading to the development of imaging agents 64Cu-DOTA- ZHER2:477 and 64Cu-DOTA-(ZHER2:477)2 (9). Interestingly, these studies showed that smaller affibody constructs performed better in vivo in terms of tumor uptake and clearance in spite of the lower affinity in vitro. The investigators then generated a class of small proteins consisting of two α-helix bundles of the 3-helix affibody by deleting the helix 3 because the binding domain localizes in the α-helices 1 and 2 bundles (5). One of these 2-helix proteins is MUT-DS, which has α-helices 1 and 2 bundles, with a disulfide bridge being formed between the two inserted homocysteines (10-12). MUT-DS showed a binding affinity to HER2 in the low-nM range. The radiolabeled MUT-DS derivatives exhibited favorable pharmacokinetics for both imaging and therapy of HER2-expressing tumors (refer to MUT-DS derived agents in MICAD). This series of chapters summarizes the data obtained with the ZHER2:477 derivatives, and this chapter presents the data obtained with 18F-FBO-ZHER2:477 (1).
Sections
Identification of Putative Molecular Imaging Probes for BACE-1 by Accounting for Protein Flexibility in Virtual Screening.
Source
Center for Molecular Medicine, School of Life Science and Biotechnology, Dalian University of Technology, Dalian, PR China.
Abstract
β-secretase (BACE-1), an enzyme critical in the process of amyloid-β (Aβ) peptides deposition in human brain, is closely associated with the onset and progression of Alzheimer's disease (AD). A strong need exists, therefore, to identify molecular imaging probes homing at BACE-1 for use with positron emission tomography (PET) that is recognized as an effective tool for detecting AD. Through this imaging, an early diagnosis of AD could be made. Herein, to identify suitable molecular probes for use with PET, we searched the Molecular Imaging and Contrast AgentDatabase (MICAD), an online database warehousing scientific information regarding molecular imaging and contrast agents, and applied a virtual screening approach against the different confirmations of BACE-1 obtained from the World Wide Protein Database. The lack of considering receptor flexibility is a key drawback in virtual screening for drug discovery. Therefore, we incorporated protein flexibility into the virtual screening by using an ensemble of 143 experimental BACE-1 structures derived from the Protein Data Bank. Finally, the best performing affinity was recorded and used in the ranking of each ligand. To the best of our knowledge, this is the first virtual screening approach used to identify four new molecular probes that could target BACE-1 with favorable affinity, a discovery that can lead to the development of new PET probes for the early detection and therapy of AD. However, the actual utility of these probes can only be ascertained after in vitro and in vivo investigations.
A comprehensive library of blocked dipeptides reveals intrinsic backbone conformational propensities of unfolded proteins.
Source
Department of Chemistry, Korea University, Seoul 136-701, Korea.
Abstract
Despite prolonged scientific efforts to elucidate the intrinsic peptide backbone preferences of amino-acids based on understanding of intermolecular forces, many open questions remain, particularly concerning neighboring peptideinteraction effects on the backbone conformational distribution of short peptides and unfolded proteins. Here, we show that spectroscopic studies of a complete library of 400 dipeptides reveal that, irrespective of side-chain properties, the backbone conformation distribution is narrow and they adopt polyproline II and β-strand, indicating the importance of backbone peptide solvation and electronic effects. By directly comparing the dipeptide circular dichroism and NMR results with those of unfolded proteins, the comprehensive dipeptides form a complete set of structural motifs of unfolded proteins. We thus anticipate that the present dipeptide library with spectroscopic data can serve as a useful databasefor understanding the nature of unfolded protein structures and for further refinements of molecular mechanical parameters. Proteins 2011; © 2011 Wiley Periodicals, Inc.
Copyright © 2011 Wiley Periodicals, Inc.
Convenient nomenclature of cysteine-rich polypeptide toxins from sea anemones.
Source
Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, 117997 Moscow, Russia.
Abstract
Polypeptide toxins are the main constituents of natural venoms. Considerable progress in the study of these molecules has resulted in the determination of a large number of structurally related sequences. To classify newly discovered molecules, a rational nomenclature for naming peptide toxins was developed, which takes into account toxin biological activity, the species name, and structural peculiarities of the polypeptide. Herein, we suggest modifications to this nomenclature for cysteine-rich polypeptide toxins from sea anemones and describe 11 novel polypeptide structures deduced after common database revision.
Copyright © 2011. Published by Elsevier Inc.
Fluorinated and iodinated (Z)-2-(4-(2-fluoroethoxy)benzylidene)-5-iodobenzofuran-3(2H)-one.
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Nov 30 [updated 2011 Dec 28].
Excerpt
Fluorinated and iodinated (Z)-2-(4-(2-fluoroethoxy)benzylidene)-5-iodobenzofuran-3(2H)-one (compound 3), abbreviated as [18F]3 and [125I]3, respectively, is an aurone derivative synthesized by Watanabe et al. for single-photon emission computed tomography (SPECT) and positron emission tomography (PET) of Alzheimer’s disease (AD) by targeting β-amyloid (Aβ) plaques (1). AD is characterized in pathology by the presence of extracellular Aβ plaques, intraneuronal neurofibrillary tangles, and neuronal loss in the cerebral cortex (2, 3). Of them, Aβ deposit is the earliest neuropathological marker and is relatively specific to AD and closely related disorders. Aβ plaques are composed of abnormal paired helical filaments 5–10 nm in size. These filaments are largely made of insoluble Aβ peptides that are 40 or 42 amino acids in length (4). In recent years, molecular imaging by targeting the extracellular Aβ has been intensively investigated in attempts to detect early AD, assess Aβ content in vivo, determine the timing of anti-plaque therapy, and evaluate the therapeutic efficacy (4). Radiolabeled Aβ40 peptides were tested first, but they showed poor penetration ability to cross the blood–brain barrier (BBB) (4). Based on the fact that Aβ can be specifically stained in vitro with dyes of Congo red, chrysamine G, and thioflavin-T, an effort was made to develop imaging agents with these dyes. This effort, however, was in general unsuccessful because the bulky ionic groups of heteroatoms in these dyes prevent them from crossing the BBB (2). Importantly, a large class of derivatives (e.g., aminonaphthalenes, benzothiazoles, stilbenes, and imidazopyridines) was synthesized with these dyes as templates (4). Clinical and preclinical studies have shown that these derivatives not only possess a high binding affinity with Aβ plaques as their parent compounds, but also exhibit good penetration ability through the BBB and rapid washout from brain. Ono et al. first synthesized a class of radioiodinated flavone derivatives that present a high binding affinity with Aβ plaques and good penetration ability through the BBB (5). However, these flavone derivatives display poor clearance from the brain, which leads to a high brain background. The investigators then explored another class of flavonoids with aurone as the core structure (6, 7). Aurone is a heterocyclic chemical compound that contains a benzofuran element associated with a benzylidene linked in position 2 and a chalcone-like group being closed into a five-member ring. The aurone derivatives possess a nucleophilic group (NH2, NHMe, or NMe2) at the 4' position and a radioiodine at the 5 position. Although these aurone derivatives exhibit a strong binding affinity with Aβ (inhibition constant (K i) = 1.2–6.8 nM), high penetration ability through the BBB (1.9%−4.6% injected dose per gram tissue (ID/g) at 2 min), and a fast washout from the brain (0.3%−0.5% ID/g at 30 min), the pharmacokinetics of these compounds are less favorable for brain imaging than the pharmacokinetics of the agent [123I]IMPY (6-iodo-2-(4'-dimethylamino)phenyl-imidazo[1,2]pyridine), which is the only SPECT agent to be tested in humans to date (1, 8, 9). The investigators also modified the flavone and aurone derivatives by pegylating them with 1–3 units of ethylene glycol at the 4' position or by conjugating them with the chelating agent bis-amino-bis-thiol (BAT) (7). Favorable pharmacokinetics for brain imaging was observed for the pegylated derivatives ([18F]8(a–c)) but not for the BAT-chelated derivatives ([99mTc]BAT-FL and [99mTc]BAT-AR) (6, 7). This series of chapters summarizes the data obtained with flavone and aurone derivatives, including [125I]15, [125I]9, [125I]14, [125I]16, [125I]17, [99mTc]BAT-FL, [99mTc]BAT-AR, [18F]8(a-c), [125I]3, and [18F]3 (1, 6-8). This chapter presents the data obtained with [125I]3 and [18F]3 (1).
Sections
Radioiodinated (Z)-2-(4-(2-hydroxyethoxy)benzylidene)-5-iodobenzofuran-3(2H)-one.
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Nov 30 [updated 2011 Dec 28].
Excerpt
Radioiodinated (Z)-2-(4-(2-hydroxyethoxy)benzylidene)-5-iodobenzofuran-3(2H)-one (compound 15), abbreviated as [125I]15, is an aurone derivative synthesized by Maya et al. for single-photon emission computed tomography(SPECT) of Alzheimer’s disease (AD) by targeting β-amyloid (Aβ) (1). The other four aurone derivatives include radioiodinated (Z)-2-(4-methoxybenzylidene)-5-iodobenzofuran-3(2H)-one (compound 9), (Z)-2-(4-hydroxybenzylidene)-5-iodobenzofuran-3(2H)-one (compound 14), (Z)-2-(4-(2-(2-hydroxyethoxy)ethoxy)benzylidene)-5-iodobenzofuran-3(2H)-one (compound 16), and (Z)-2-(4-(2-(2-(2-hydroxyethoxy)ethoxy)ethoxy)benzylidene)-5-iodobenzofuran-3(2H)-one (compound 17), which are abbreviated as [125I]9, [125I]14, [125I]16, and [125I]17, respectively. AD is characterized in pathology by the presence of extracellular Aβ plaques, intraneuronal neurofibrillary tangles, and neuronal loss in the cerebral cortex (2, 3). Of them, Aβ deposit is the earliest neuropathological marker and is relatively specific to AD and closely related disorders. Aβ plaques are composed of abnormal paired helical filaments 5–10 nm in size. These filaments are largely made of insoluble Aβ peptides that are 40 or 42 amino acids in length (4). In recent years, molecular imaging by targeting the extracellular Aβ has been intensively investigated in attempts to detect early AD, assess Aβ content in vivo, determine the timing of anti-plaque therapy, and evaluate the therapeutic efficacy (4). Radiolabeled Aβ40 peptides were tested first, but they showed poor penetration ability to cross the blood–brain barrier (BBB) (4). Based on the fact that Aβ can be specifically stained in vitro with dyes of Congo red, chrysamine G, and thioflavin-T, an effort was made to develop imaging agents with these dyes. This effort, however, was in general unsuccessful because the bulky ionic groups of heteroatoms in these dyes prevent them from crossing the BBB (2). Importantly, a large class of derivatives (e.g., aminonaphthalenes, benzothiazoles, stilbenes, and imidazopyridines) was synthesized with these dyes as templates (4). Clinical and preclinical studies have shown that these derivatives not only possess a high binding affinity with Aβ plaques as their parent compounds, but also exhibit good penetration ability through the BBB and rapid washout from brain with low to no plaque deposits. Ono et al. first synthesized a class of radioiodinated flavone derivatives that present a high binding affinity with Aβ plaques and good penetration ability through the BBB (5). However, these flavone derivatives display poor clearance from the brain, which leads to a high brain background. The investigators then explored another class of flavonoids with aurone as the core structure (6, 7). Aurone is a heterocyclic chemical compound that contains a benzofuran element associated with a benzylidene linked in position 2 and a chalcone-like group being closed into a five-member ring. The aurone derivatives possess a nucleophilic group (NH2, NHMe, or NMe2) at the 4' position and a radioiodine at the 5 position. Although these aurone derivatives exhibit a strong binding affinity with Aβ (inhibition constant (K i) = 1.2–6.8 nM), high penetration ability through the BBB (1.9%−4.6% injected dose per gram tissue (ID/g) at 2 min), and a fast washout from the brain (0.3%−0.5% ID/g at 30 min), the pharmacokinetics of these compounds are less favorable for brain imaging than the pharmacokinetics of the agent [123I]IMPY (6-iodo-2-(4'-dimethylamino)phenyl-imidazo[1,2]pyridine), which is the only SPECT agent to be tested in humans to date (1, 8, 9). The investigators also modified the flavone and aurone derivatives by pegylating them with 1–3 units of ethylene glycol at the 4' position or by conjugating them with the chelating agent bis-amino-bis-thiol (BAT) (7). Favorable pharmacokinetics for brain imaging was observed for the pegylated derivatives ([18F]8(a–c)) but not for the BAT-chelated derivatives ([99mTc]BAT-FL and [99mTc]BAT-AR) (6, 7). This series of chapters summarizes the data obtained with flavone and aurone derivatives, including [125I]15, [125I]9, [125I]14, [125I]16, [125I]17, [99mTc]BAT-FL, [99mTc]BAT-AR, [18F]8(a–c), [125I]3, and [18F]3 (1, 6-8). This chapter presents the data obtained with [125I]15, [125I]9, [125I]14, [125I]16, and [125I]17 (1).
Sections
99mTc-Bis-amino-bis-thiol-conjugated 6-(3-bromopropoxy)-2-(4-(dimethylamino)phenyl)-4H-chromen-4-one and (Z)-5-(3-bromopropoxy)-2-(4-(dimethylamino)benzylidene)benzofuran-3(2H)-one.
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Nov 30 [updated 2011 Dec 28].
Excerpt
99mTc-Bis-amino-bis-thiol (BAT)-conjugated 6-(3-bromopropoxy)-2-(4-(dimethylamino)phenyl)-4H-chromen-4-one and (Z)-5-(3-bromopropoxy)-2-(4-(dimethylamino)benzylidene)benzofuran-3(2H)-one, abbreviated as [99mTc]BAT-FL and [99mTc]BAT-AR, respectively, are flavone and aurone derivatives synthesized by Ono et al. for single-photon emission computed tomography (SPECT) of Alzheimer’s disease (AD) by targeting β-amyloid (Aβ) (1). AD is characterized in pathology by the presence of extracellular Aβ plaques, intraneuronal neurofibrillary tangles, and neuronal loss in the cerebral cortex (2, 3). Of them, Aβ deposit is the earliest neuropathological marker and is relatively specific to AD and closely related disorders. Aβ plaques are composed of abnormal paired helical filaments 5–10 nm in size. These filaments are largely made of insoluble Aβ peptides that are 40 or 42 amino acids in length (4). In recent years, molecular imaging by targeting the extracellular Aβ has been intensively investigated in attempts to detect early AD, assess Aβ content in vivo, determine the timing of anti-plaque therapy, and evaluate the therapeutic efficacy (4). Radiolabeled Aβ40 peptides were tested first, but they showed poor penetration ability to cross the blood–brain barrier (BBB) (4). Based on the fact that Aβ can be specifically stained in vitro with dyes of Congo red, chrysamine G, and thioflavin-T, an effort was made to develop imaging agents with these dyes. This effort, however, was in general unsuccessful because the bulky ionic groups of heteroatoms in these dyes prevent them from crossing the BBB (2). Importantly, a large class of derivatives (e.g., aminonaphthalenes, benzothiazoles, stilbenes, and imidazopyridines) was synthesized with these dyes as templates (4). Clinical and preclinical studies have shown that these derivatives not only possess a high binding affinity with Aβ plaques as their parent compounds, but also exhibit good penetration ability through the BBB and rapid washout from brain with low to no plaque deposits. Ono et al. first synthesized a class of radioiodinated flavone derivatives that present a high binding affinity with Aβ plaques and good penetration ability through the BBB (5). However, these flavone derivatives display poor clearance from the brain, which leads to a high brain background. The investigators then explored another class of flavonoids with aurone as the core structure (1, 6). Aurone is a heterocyclic chemical compound that contains a benzofuran element associated with a benzylidene linked in position 2 and a chalcone-like group closed into a five-member ring. The aurone derivatives possess a nucleophilic group (NH2, NHMe, or NMe2) at the 4' position and a radioiodine at the 5 position. Although these aurone derivatives exhibit a strong binding affinity with Aβ (inhibition constant (K i) = 1.2–6.8 nM), high penetration ability through the BBB (1.9%−4.6% injected dose per gram tissue (ID/g) at 2 min), and a fast washout from the brain (0.3%−0.5% ID/g at 30 min), the pharmacokinetics of these compounds are less favorable for brain imaging than the pharmacokinetics of the agent [123I]IMPY (6-iodo-2-(4'-dimethylamino)phenyl-imidazo[1,2]pyridine), which is the only SPECT agent to be tested in humans to date (7-9). The investigators also modified the flavone and aurone derivatives by pegylating them with 1–3 units of ethylene glycol at the 4' position or by conjugating them with the chelating agent bis-amino-bis-thiol (BAT). Favorable pharmacokinetics for brain imaging was observed for the pegylated derivatives ([18F]8(a–c)) but not for the BAT-chelated derivatives ([99mTc]BAT-FL and [99mTc]BAT-AR) (1, 6). This series of chapters summarizes the data obtained with flavone and aurone derivatives, including [125I]15, [125I]9, [125I]14, [125I]16, [125I]17, [99mTc]BAT-FL, [99mTc]BAT-AR, [18F]8(a–c), [125I]3, and [18F]3 (1, 6-8). This chapter presents the data obtained with [99mTc]BAT-FL and [99mTc]BAT-AR (1).
Sections
18F-Labeled fluoropegylated 6-fluoroethoxy-4'-dimethylaminoflavone, 6-(2-(2-fluoro-ethoxy)-ethoxy)-4'-dimethylaminoflavone, and 6-(2-(2-(2-fluoro-ethoxy)-ethoxy)ethoxy)-4'-dimethylaminoflavone.
Authors
Source
Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2011.
2011 Nov 30 [updated 2011 Dec 28].
Excerpt
18F-Labeled fluoropegylated 6-fluoroethoxy-4'-dimethylaminoflavone (compound 8a), 6-(2-(2-fluoro-ethoxy)-ethoxy)-4'-dimethylaminoflavone (compound 8b), and 6-(2-(2-(2-fluoro-ethoxy)-ethoxy)ethoxy)-4'-dimethylaminoflavone (compound 8c), abbreviated as [18F]8a, [18F]8b, and [18F]8c, respectively, are flavone derivatives synthesized by Ono et al. for positron emission tomography (PET) of Alzheimer’s disease (AD) by targeting β-amyloid (Aβ) (1). AD is characterized in pathology by the presence of extracellular Aβ plaques, intraneuronal neurofibrillary tangles, and neuronal loss in the cerebral cortex (2, 3). Of them, Aβ deposit is the earliest neuropathological marker and is relatively specific to AD and closely related disorders. Aβ plaques are composed of abnormal paired helical filaments 5–10 nm in size. These filaments are largely made of insoluble Aβ peptides that are 40 or 42 amino acids in length (4). In recent years, molecular imaging by targeting the extracellular Aβ has been intensively investigated in attempts to detect early AD, assess Aβ content in vivo, determine the timing of anti-plaque therapy, and evaluate the therapeutic efficacy (4). Radiolabeled Aβ40 peptides were tested first, but they showed poor penetration ability to cross the blood–brain barrier (BBB) (4). Based on the fact that Aβ can be specifically stained in vitro with dyes of Congo red, chrysamine G, and thioflavin-T, an effort was made to develop imaging agents with these dyes. This effort, however, was in general unsuccessful because the bulky ionic groups of heteroatoms in these dyes prevent them from crossing the BBB (2). Importantly, a large class of derivatives (e.g., aminonaphthalenes, benzothiazoles, stilbenes, and imidazopyridines) was synthesized with these dyes as templates (4). Clinical and preclinical studies have shown that these derivatives not only possess a high binding affinity with Aβ plaques as their parent compounds, but also exhibit good penetration ability through the BBB and rapid washout from brain with low to no plaque deposits. Ono et al. first synthesized a class of radioiodinated flavone derivatives that present a high binding affinity with Aβ plaques and good penetration ability through the BBB (5). However, these flavone derivatives display poor clearance from the brain, which leads to a high brain background. The investigators then explored another class of flavonoids with aurone as the core structure (1, 6). Aurone is a heterocyclic chemical compound that contains a benzofuran element associated with a benzylidene linked in position 2 and a chalcone-like group being closed into a five-member ring. The aurone derivatives possess a nucleophilic group (NH2, NHMe, or NMe2) at the 4' position and a radioiodine at the 5 position. Although these aurone derivatives exhibit a strong binding affinity with Aβ (inhibition constant (K i) = 1.2–6.8 nM), high penetration ability through the BBB (1.9%−4.6% injected dose per gram tissue (ID/g) at 2 min), and a fast washout from the brain (0.3%−0.5% ID/g at 30 min), the pharmacokinetics of these compounds are less favorable for brain imaging than the pharmacokinetics of the agent [123I]IMPY (6-iodo-2-(4'-dimethylamino)phenyl-imidazo[1,2]pyridine), which is the only SPECT agent to be tested in humans to date (7-9). The investigators also modified the flavone and aurone derivatives by pegylating them with 1–3 units of ethylene glycol at the 4' position or by conjugating them with the chelating agent bis-amino-bis-thiol (BAT). Favorable pharmacokinetics for brain imaging was observed for the pegylated derivatives ([18F]8(a–c)) but not for the BAT-chelated derivatives ([99mTc]BAT-FL and [99mTc]BAT-AR) (1, 6). This series of chapters summarizes the data obtained with flavone and aurone derivatives, including [125I]15, [125I]9, [125I]14, [125I]16, [125I]17, [99mTc]BAT-FL, [99mTc]BAT-AR, [18F]8(a–c), [125I]3, and [18F]3 (1, 6-8). This chapter presents the data obtained with [18F]8(a–c) (1).
Sections
Pepitome: evaluating improved spectral library search for identification complementarity and quality assessment.
Abstract
Spectral libraries have emerged as a viable alternative to protein sequence databases for peptide identification. These libraries contain previously detected peptide sequences and their corresponding tandem mass spectra (MS/MS). Search engines can then identify peptides by comparing experimental MS/MS scans to those in the library. Many of these algorithms employ the dot product score for measuring the quality of a spectrum-spectrum match (SSM). This scoring system does not offer a clear statistical interpretation and ignores fragment ion m/z discrepancies in the scoring. We developed a new spectral library search engine, Pepitome, which employs statistical systems for scoring SSMs. Pepitome outperformed the leading library search tool, SpectraST, when analyzing data sets acquired on three different mass spectrometry platforms. We characterized the reliability of spectral library searches by confirming shotgun proteomics identifications through RNA-Seq data. Applying spectral library and database searches on the same sample revealed their complementary nature. Pepitome identifications enabled the automation of quality analysis and quality control (QA/QC) for shotgun proteomics data acquisition pipelines.
BuildSummary: Using group-based approach to improve the sensitivity ofpeptide/protein identification in shotgun proteomics.
Abstract
The target-decoy database search strategy is widely accepted as a standard method for estimating the false discovery rate (FDR) of peptide identification, based on which peptide-spectrum matches (PSMs) from the target database are filtered. To improve the sensitivity of protein identification given a fixed accuracy (frequently defined by a protein FDR threshold), a post-processing procedure is often used that integrates results from different peptide search engines that had assayed the same dataset. In this work, we show that PSMs that are grouped by the precursor charge, the number of missed internal cleavage sites, the modification state, the numbers of protease termini and the proteins grouped by their unique peptide count should be filtered separately according to the given FDR. We also develop an iterative procedure to filter the PSMs and proteins simultaneously, according to the given FDR. Finally, we present a general framework to integrate the results from different peptide search engines using the same FDR threshold. Our method was tested with several shotgun proteomics datasets that were acquired by multiple LC/MS instruments from two different biological samples. The results showed a satisfactory performance. We implemented the method in a user-friendly software package called BuildSummary, which can be downloaded for free from http://www.proteomics.ac.cn/software/proteomicstools/index.htm as part of the software suite ProteomicsTools.
GuiTope: An Application for Mapping Random-Sequence Peptides to Protein Sequences.
Abstract
ABSTRACT:
BACKGROUND:
Random-sequence peptide libraries are a commonly used tool to identify novel ligands for binding antibodies, other proteins, and small molecules. It is often of interest to compare the selected peptide sequences to the natural protein binding partners to infer the exact binding site or the importance of particular residues. The ability to search a set of sequences for similarity to a set of peptides may sometimes enable the prediction of an antibody epitope or a novel binding partner. We have developed a software application designed specifically for this task.
RESULTS:
GuiTope provides a graphical user interface for aligning peptide sequences to protein sequences. All alignment parameters are accessible to the user including the ability to specify the underlying amino acid frequency in the peptide library, which often differs significantly from the frequencies assumed by popular alignment programs. It also includes a novel feature to align di-peptide inversions, which we have found improves the accuracy of antibody epitope prediction from peptide microarray data and shows utility in analyzing phage display datasets. Finally, GuiTope can randomly select peptides from a given library to estimate a null distribution of scores to calculate statistical significance.
CONCLUSIONS:
GuiTope provides a convenient method for comparing selected peptide sequences to protein sequences, including flexible alignment parameters, novel alignment features, ability to search a database, and statistical significance of results. The latest version of the software available as an executable (for PC) at www.immunosignature.com/software and ongoing updates and source code will be available at sourceforge.net.
An enhanced protein crosslink identification strategy using CID-cleavable chemical crosslinkers and LC/MS(n) analysis.
Source
Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC, USA.
Abstract
We describe a novel two-step LC/MS(n) strategy to effectively and confidently identify numerous crosslinked peptidesfrom complex mixtures. This method incorporates the use of our gas-phase cleavable crosslinking reagent, disuccinimidyl-succinamyl-aspartyl-proline (SuDP), and a new data-processing algorithm CXLinkS (Cleavable Crosslink Selection), which enables unequivocal crosslink peptide selection and identification on the basis of mass measurement accuracy, high resolving power, and the unique fragmentation pattern of each crosslinked peptide. We demonstrate our approach with well-characterized monomeric and multimeric protein systems with and without database searching restrictions where inter-peptide crosslink identification is increased 8-fold over our previously published data-dependent LC/MS(3) method and discuss its applicability to other CID-cleavable crosslinkers and more complex protein systems.
Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Biomonitors of cardiac injury and performance: B-type natriuretic peptide and troponin as monitors of hemodynamics and oxygen transport balance.
Source
Children's Hospital of Orange County, Orange, CA, USA.
Abstract
Serum biomarkers, such as B-type natriuretic peptide and troponin, are frequently measured in the cardiac intensive care unit. A review of the evidence supporting monitoring of these biomarkers is presented.
DESIGN:
A search of MEDLINE, PubMed, and the Cochrane Database was conducted to find literature regarding the use of B-type natriuretic peptide and troponin in the cardiac intensive care setting. Adult and pediatric data were considered.
RESULTS AND CONCLUSION:
Both B-type natriuretic peptide and troponin have demonstrated utility in the intensive care setting but there is no conclusive evidence at this time that either biomarker can be used to guide inpatient management of children with cardiac disease. Although B-type natriuretic peptide and troponin concentrations can alert clinicians to myocardial stress, injury, or hemodynamic alterations, the levels can also be elevated in a variety of clinical scenarios, including sepsis. Observational studies have demonstrated that perioperative measurement of these biomarkers can predict postoperative mortality and complications. RECOMMENDATION AND
LEVEL OF EVIDENCE:
(class IIb, level of evidence B): The use of B-type natriuretic peptide and/or troponin measurements in the evaluation of hemodynamics and postoperative outcome in pediatric cardiac patients may be beneficial.
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