Publications by authors named "Marcin Krajewski"

21 Publications

  • Page 1 of 1

Enhancement of mechanical properties of vertically aligned carbon nanotube arrays due to N ion irradiation.

Nanotechnology 2020 Apr 3;31(28):285703. Epub 2020 Apr 3.

Institute of Fundamental Technological Research, Polish Academy of Sciences, Warsaw, Poland. Warsaw University of Technology, Faculty of Mechatronics, Warsaw, Poland.

In this work we apply N ion irradiation on vertically aligned carbon nanotube (VACNT) arrays in order to increase the number of connections and joints in the CNT network. The ions energy was 50 keV and fluence 5 × 10 ions cm. The film was 160 μm thick. SEM images revealed the ion irradiation altered the carbon bonding and created a sponge-like, brittle structure at the surface of the film, with the ion irradiation damage region extending ∼4 μm in depth. TEM images showed the brittle structure consists of amorphous carbon forming between nanotubes. The significant enhancement of mechanical properties of the irradiated sample studied by the cyclic nanoindentation with a flat punch indenter was observed. Irradiation on the VACNT film made the structure stiffer, resulted in a higher percentage recovery, and reduced the energy dissipation under compression. The results are encouraging for further studies which will lead to create a class of materials-ion-irradiated VACNT films-which after further research may find application in storage or harvesting energy at the micro/nanoscale.
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http://dx.doi.org/10.1088/1361-6528/ab8665DOI Listing
April 2020

Preliminary Studies on Biodegradable Zinc Oxide Nanoparticles Doped with Fe as a Potential Form of Iron Delivery to the Living Organism.

Nanoscale Res Lett 2019 Dec 10;14(1):373. Epub 2019 Dec 10.

Department of Physiological Sciences, Faculty of Veterinary Medicine, Warsaw University of Life Sciences, Nowoursynowska 159, 02-776, Warsaw, Poland.

Iron is the crucial element for living organisms and its deficiency is described as the most common nutritional disorder all over the world. Nowadays, more effective and safe iron supplementation strategies for both humans and animals become one of the most important challenges in the therapy of nutritional deficiencies. Our previous in vivo studies confirmed safety and biodegradability of in-house manufactured zinc oxide-based nanoparticles and their rapid distribution to majority of organs and tissues in the body. In vitro examinations performed on Caco-2 cell line, a model of epithelial cells of the gastrointestinal tract, revealed a low toxicity of studied nanomaterials. In the current study, we investigated biodegradable zinc oxide nanoparticles doped with Fe(III) as a perspective supplementation strategy for iron deficiency. Biodegradable ZnO:Fe nanoparticles were intra-gastrically administered to adult mice and following 24 h, animals were sacrificed with collection of internal organs for further analyses. The iron concentration measured with atomic absorption spectrometry and histological staining (Perl's method) showed a rapid distribution of iron-doped nanoparticles to tissues specifically related with iron homeostasis. Accumulation of iron was also visible within hepatocytes and around blood vessels within the spleen, which might indicate the transfer of Fe-doped nanoparticles from the bloodstream into the tissue. Reassuming, preliminary results obtained in the current study suggest that biodegradable ZnO nanoparticles doped with Fe might be a good carriers of exogenous iron in the living body. Therefore, subsequent investigations focus on determination an exact mechanisms related with an iron deposition in the tissue and influence of nanoparticle carriers on iron metabolism are required.
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http://dx.doi.org/10.1186/s11671-019-3217-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6904721PMC
December 2019

Magnetic-field-induced synthesis of magnetic wire-like micro- and nanostructures.

Authors:
Marcin Krajewski

Nanoscale 2017 Nov;9(43):16511-16545

Institute of Fundamental Technological Research, Polish Academy of Sciences, Pawinskiego Street 5B, 02-106 Warsaw, Poland.

A lot of physical and chemical preparation methods of one-dimensional (1D) structures are known today. Most of them use highly advanced technology or quite complex chemical reagents. This results in their high costs and difficulties with their implementation to a large industrial scale. Hence, new, facile and inexpensive approaches are still sought. One alternative to wire-like structure production is based on the chemical reduction reactions combined with an external magnetic field, which acts as an independent synthesis parameter. This approach is commonly called magnetic-field-assisted (MFA) synthesis or magnetic-field-induced (MFI) synthesis. As usual, this manufacturing strategy comprises both drawbacks and advantages, which are introduced in this review. Moreover, this work shows that MFI synthesis depends on several synthesis parameters including the strength of the applied magnetic field, reaction temperature, pH value of the reaction environment, chemical composition of the precursor solution, reaction time, and also the presence of surfactants, complexing agents, nucleating agents, initiators as well as organic solvents. All of them have an impact on the morphology and dimensions of wire-like materials and their chemical, physical and mechanical properties. Finally, the opportunities and challenges associated with the magnetic-assisted fabrication of wire-like structures are widely discussed in this review.
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http://dx.doi.org/10.1039/c7nr05823cDOI Listing
November 2017

A multisample dissolution dynamic nuclear polarization system for serial injections in small animals.

Magn Reson Med 2017 02 22;77(2):904-910. Epub 2016 Feb 22.

Institute for Biomedical Engineering, University and ETH Zurich, Switzerland.

Purpose: Several in vivo applications of dissolution dynamic nuclear polarization (DNP) require rapid successive injections of hyperpolarized substrates. Here we present the design and performance of a custom-built multisample dissolution DNP setup for small animal research.

Methods: The DNP setup consists of a commercial wide-bore magnet charged to 3.35 T, a cryostat, a 94-GHz microwave source, and a custom-built skeleton that accommodates four identical sample sticks. Each sample stick features a dissolver locked into the skeleton port and a lifter, which permits moving the sample cup out of the liquid helium bath for dissolution.

Results: The dissolution of the first sample was triggered after 2 hours of polarization buildup during single-shot operation of the cryostat. Thereafter, a time window of 75-90 min was available to dissolve the remaining three polarized samples. The average liquid state polarization over all four sticks was measured as 18.7% ± 2.3% for [1-13C] pyruvate 30 s after dissolution. In vivo applicability of the setup using serial injections of [1-13C] pyruvate to study cardiac metabolism in rats revealed good reproducibility.

Conclusion: The proposed four-sample DNP insert provides reproducible liquid state polarization of [1-13C] pyruvate and allows for rapid repeat injections in small animals. Magn Reson Med 77:904-910, 2017. © 2016 International Society for Magnetic Resonance in Medicine.
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http://dx.doi.org/10.1002/mrm.26147DOI Listing
February 2017

Hyperpolarized Metabolic MR Imaging of Acute Myocardial Changes and Recovery after Ischemia-Reperfusion in a Small-Animal Model.

Radiology 2016 Mar 23;278(3):742-51. Epub 2015 Nov 23.

From the Institute for Biomedical Engineering, University and ETH Zurich, Gloriastrasse 35, 8092 Zurich, Switzerland (D.O., P.W., J.B., L.W., M.K., K.W., A.S., S.K.); Departments of Congenital Heart Disease and Pediatric Cardiology (D.O., D.M.) and Internal Medicine, Division of Cardiology (D.M.), German Heart Institute, Berlin, Germany; and Division of Imaging Sciences and Biomedical Engineering, King's College London, London, England (D.O., S.K.).

Purpose: To implement hyperpolarized magnetic resonance (MR) imaging in an animal model of ischemia-reperfusion and to assess in vivo the regional changes in pyruvate metabolism within the 1st hour and at 1 week after a brief episode of coronary occlusion and reperfusion.

Materials And Methods: All animal experiments were performed with adherence to the Swiss Animal Protection law and were approved by the regional veterinary office. A closed-chest rat model was implemented by using an inflatable balloon secured around the left coronary artery. Animals were placed in an MR system 5-7 days after surgery. [1-(13)C]pyruvate was polarized by using a home-built multisample hyperpolarizer. Hyperpolarized pyruvate was injected at five stages: at baseline; at reperfusion after 15 minutes of coronary occlusion; and at 30 minutes, 60 minutes, and 1 week after ischemia reperfusion. The conversion of pyruvate into lactate and bicarbonate was imaged by using dedicated MR sequences alongside wall motion and delayed enhancement imaging. After imaging, the heart was removed and stained to delineate the area at risk (AAR). Differences between AAR and remote myocardium were assessed by using a repeated measures analysis of variance and a post hoc Bonferroni multiple comparison test.

Results: Data were collected in 12 animals. Occlusion led to hypokinesia of the anterior or anterolateral segments of the myocardium. At reperfusion, the average lactate-to-bicarbonate ratio increased in the AAR relative to that at baseline (from 1.93 ± 0.48 to 3.01 ± 0.74, P < .001) and was significantly higher when compared with that in the remote area (1.91 ± 0.38, P < .001). In the 60 minutes after occlusion, the lactate-to-bicarbonate ratio in the AAR recovered but was still elevated relative to that in the remote area. One week after ischemia-reperfusion, no difference between AAR and remote area could be detected.

Conclusion: Hyperpolarized metabolic MR imaging can be used to successfully detect acute changes in [1-(13)C]pyruvate metabolism after ischemia-reperfusion, thereby enabling in vivo monitoring of the metabolic effects of reperfusion strategies.
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http://dx.doi.org/10.1148/radiol.2015151332DOI Listing
March 2016

Structural and magnetic properties of iron nanowires and iron nanoparticles fabricated through a reduction reaction.

Beilstein J Nanotechnol 2015 29;6:1652-60. Epub 2015 Jul 29.

University of Warsaw, Faculty of Physics, Institute of Experimental Physics, Pasteura St. 5, 02-093 Warsaw, Poland.

The main goal of this work is to study the structural and magnetic properties of iron nanowires and iron nanoparticles, which have been fabricated in almost the same processes. The only difference in the synthesis is an application of an external magnetic field in order to form the iron nanowires. Both nanomaterials have been examined by means of transmission electron microscopy, energy dispersive X-ray spectrometry, X-ray diffractometry and Mössbauer spectrometry to determine their structures. Structural investigations confirm that obtained iron nanowires as well as nanoparticles reveal core-shell structures and they are composed of crystalline iron cores that are covered by amorphous or highly defected phases of iron and iron oxides. Magnetic properties have been measured using a vibrating sample magnetometer. The obtained values of coercivity, remanent magnetization, saturation magnetization as well as Curie temperature differ for both studied nanostructures. Higher values of magnetizations are observed for iron nanowires. At the same time, coercivity and Curie temperature are higher for iron nanoparticles.
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http://dx.doi.org/10.3762/bjnano.6.167DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4578357PMC
October 2015

Hybrid multiband excitation multiecho acquisition for hyperpolarized (13) C spectroscopic imaging.

Magn Reson Med 2015 May 20;73(5):1713-7. Epub 2014 May 20.

Institute for Biomedical Engineering, University and ETH Zurich, Zurich, Switzerland.

Purpose: Fast dynamic imaging of hyperpolarized (13) C-labeled pyruvate and its downstream metabolites shows great potential for probing metabolic changes in the heart. Sequences that allow for fast encoding of the spectral and spatial information of the myocardial metabolism and optimal signal excitation are usually limited by gradient performance, especially at high magnetic fields. Here we propose a combination of a spectral-spatial multiband excitation and multiecho readout to overcome these limitations.

Methods: By using a low-bandwidth, two-pulse excitation, a thinner slice compared with conventional spectral-spatial excitation is achieved, while at the same time allowing for low flip angle excitation on pyruvate and high flip angle excitation on bicarbonate and lactate, which optimizes signal-to-noise ratio (SNR) in cardiac metabolic imaging. The implementation was evaluated in 13 healthy female Sprague-Dawley rats at 9.4T.

Results: Using a slice thickness of 4 mm, a mean (± standard deviation) peak SNR of 18.3 (±8.4), 15.2 (±6.6), and 8.6 (±2.0) was observed for pyruvate, lactate, and bicarbonate, respectively.

Conclusion: This approach provides high SNR in metabolic images while at the same time allowing for a thin slice selection even at high magnetic fields. This is crucial in metabolic imaging in small animal models.
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http://dx.doi.org/10.1002/mrm.25294DOI Listing
May 2015

Accelerating hyperpolarized metabolic imaging of the heart by exploiting spatiotemporal correlations.

NMR Biomed 2013 Nov 25;26(11):1380-6. Epub 2013 Apr 25.

Institute for Biomedical Engineering, University and ETH Zurich, Zurich, Switzerland.

Hyperpolarized (13)C-labeled pyruvate is a promising tool to investigate cardiac metabolism. It has been shown that changes in substrate metabolism occur following the induction of ischemia. To investigate the metabolic changes that are confined to spatial regions, high spatiotemporal resolution is required. The present work exploits both spatial and temporal correlations using k-t principal component analysis (PCA) to undersample the spatiotemporal domain, thereby speeding up data acquisition. A numerical model was implemented to investigate optimal acquisition and reconstruction parameters for pyruvate, lactate and bicarbonate maps of the heart. Subsequently, prospectively undersampled in vivo data on rat hearts were acquired using a combination of spectral-spatial signal excitation and a variable-density single-shot echo planar readout. Using five-fold k-t PCA, a spatial resolution of 1 × 1 mm(2) at a temporal resolution of 3 s was achieved.
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http://dx.doi.org/10.1002/nbm.2963DOI Listing
November 2013

A multi-sample 94 GHz dissolution dynamic-nuclear-polarization system.

J Magn Reson 2012 Jan 15;214(1):166-74. Epub 2011 Nov 15.

Physical Chemistry, ETH Zürich, Wolfgang-Pauli-Strasse 10, 8093 Zürich, Switzerland.

We describe the design and initial performance results of a multi-sample dissolution dynamic-nuclear-polarization (DNP) polarizer based on a Helium-temperature NMR cryostat for use in a wide-bore NMR magnet with a room-temperature bore. The system is designed to accommodate up to six samples in a revolver-style sample changer that allows changing samples at liquid-Helium temperature and at pressures ranging from ambient pressure down to 1 mbar. The multi-sample setup is motivated by the desire to do repetitive in vivo measurements and to characterize the DNP process by investigating samples of different chemical composition. The system can be loaded with up to six samples simultaneously to reduce sample loading and unloading. Therefore, series of experiments can be carried out faster and more reliably. The DNP probe contains an oversized microwave cavity and includes EPR and NMR capabilities for monitoring the DNP process. In the solid state, DNP enhancements corresponding to ∼45% polarization for [1-(13)C]pyruvic acid with a trityl radical have been measured. In the initial liquid-state acquisition experiments described here, the polarization was found to be ∼13%, corresponding to an enhancement factor exceeding 16,000 relative to thermal polarization at 9.4 T and ambient temperature.
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http://dx.doi.org/10.1016/j.jmr.2011.11.002DOI Listing
January 2012

Role of the ubiquitin-like protein Hub1 in splice-site usage and alternative splicing.

Nature 2011 May 25;474(7350):173-8. Epub 2011 May 25.

Department of Molecular Cell Biology, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.

Alternative splicing of pre-messenger RNAs diversifies gene products in eukaryotes and is guided by factors that enable spliceosomes to recognize particular splice sites. Here we report that alternative splicing of Saccharomyces cerevisiae SRC1 pre-mRNA is promoted by the conserved ubiquitin-like protein Hub1. Structural and biochemical data show that Hub1 binds non-covalently to a conserved element termed HIND, which is present in the spliceosomal protein Snu66 in yeast and mammals, and Prp38 in plants. Hub1 binding mildly alters spliceosomal protein interactions and barely affects general splicing in S. cerevisiae. However, spliceosomes that lack Hub1, or are defective in Hub1-HIND interaction, cannot use certain non-canonical 5' splice sites and are defective in alternative SRC1 splicing. Hub1 confers alternative splicing not only when bound to HIND, but also when experimentally fused to Snu66, Prp38, or even the core splicing factor Prp8. Our study indicates a novel mechanism for splice site utilization that is guided by non-covalent modification of the spliceosome by an unconventional ubiquitin-like modifier.
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http://dx.doi.org/10.1038/nature10143DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3587138PMC
May 2011

Isoquinolin-1-one inhibitors of the MDM2-p53 interaction.

ChemMedChem 2008 Jul;3(7):1118-28

Max Planck Institute for Biochemistry, 82152 Martinsried, Germany.

p53 has been at the centre of attention for drug design since the discovery of its growth-suppressive and pro-apoptotic activity. Herein we report the design and characterisation of a new class of isoquinolinone inhibitors of the MDM2-p53 interaction. Our identification of druglike and selective inhibitors of this protein-protein interaction included a straightforward in silico compound-selection process, a recently reported NMR spectroscopic approach for studying the MDM2-p53 interaction, and selectivity screening assays using cells with the same genetic background. The selected inhibitors were all able to induce apoptosis and the expression of p53-related genes, but only the isoquinolin-1-one-based inhibitors stabilised p53. Our NMR experiments give a persuading explanation for these results, showing that isoquinolin-1-one derivates are able to dissociate the preformed MDM2-p53 complex in vitro, releasing a folded and soluble p53. The joint application of these methods provides a framework for the discovery of protein interaction inhibitors as a promising starting point for further drug design.
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http://dx.doi.org/10.1002/cmdc.200800025DOI Listing
July 2008

Molecular basis for the inhibition of p53 by Mdmx.

Cell Cycle 2007 Oct 12;6(19):2386-92. Epub 2007 Oct 12.

Max Planck Institute for Biochemistry, Martinsried, Germany.

The oncoprotein Mdm2, and the recently intensely studied, homologues protein Mdmx, are principal negative regulators of the p53 tumor suppressor. The mechanisms by which they regulate the stability and activity of p53 are not fully established. We have determined the crystal structure of the N-terminal domain of Mdmx bound to a 15-residue p53 peptide. The structure reveals that although the principle features of the Mdm2-p53 interaction are preserved in the Mdmx-p53 complex, the Mdmx hydrophobic cleft on which the p53 peptide binds is significantly altered: a part of the cleft is blocked by sidechains of Met and Tyr of the p53-binding pocket of Mdmx. Thus specific inhibitors of Mdm2-p53 would not be optimal for binding to Mdmx. Our binding assays show indeed that nutlins, the newly discovered, potent antagonists of the Mdm2-p53 interaction, are not capable to efficiently disrupt the Mdmx-p53 interaction. To achieve full activation of p53 in tumor cells, compounds that are specific for Mdmx are necessary to complement the Mdm2 specific binders.
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http://dx.doi.org/10.4161/cc.6.19.4740DOI Listing
October 2007

An NMR-based antagonist induced dissociation assay for targeting the ligand-protein and protein-protein interactions in competition binding experiments.

J Med Chem 2007 Sep 16;50(18):4382-7. Epub 2007 Aug 16.

Max Planck Institute for Biochemistry, D-82152 Martinsried, Germany.

We present an NMR-based antagonist induced dissociation assay (AIDA) for validation of inhibitor action on protein-protein interactions. As opposed to many standard NMR methods, AIDA directly validates the inhibitor potency in an in vitro NMR competition binding experiment. AIDA requires a large protein fragment (larger than 30 kDa) to bind to a small reporter protein (less than 20 kDa). We show here that a small fragment of a protein fused to glutathione S-transferase (GST) can effectively substitute the large protein component. We successfully used a GST-tagged N-terminal 73-residue p53 domain for binding studies with the human MDM2 protein. Other interactions we studied involved complexes of CDK2, cyclin A, p27, and the retinoblastoma protein. All these proteins play a key role in the cell division cycle, are associated with tumorigenesis, and are thus the subject of anticancer therapy strategies.
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http://dx.doi.org/10.1021/jm070365vDOI Listing
September 2007

Structural ramification for acetyl-lysine recognition by the bromodomain of human BRG1 protein, a central ATPase of the SWI/SNF remodeling complex.

Chembiochem 2007 Jul;8(11):1308-16

Max-Planck-Institute for Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.

Bromodomains represent an extensive family of evolutionarily conserved domains that are found in many chromatin-associated proteins such as histone acetyltransferases (HAT) and subunits of ATP-dependent chromatin-remodeling complexes. These domains are associated with acetylated lysine residues that bind both in vivo and in vitro; for example, they bind to the N-acetylated lysines of the histone tail of nucleosomes. In this report, we determined the structure of the bromodomain from human brahma-related gene 1 (BRG1) protein, a subunit of an ATP-dependent switching/sucrose nonfermenting (SWI/SNF) remodeling complex, and have also characterized its in vitro interaction with N-acetylated lysine peptides from histones. In addition to a typical all-alpha-helical fold that was observed in the bromodomains, we observed for the first time a small beta-sheet in the ZA loop region of the BRG1 protein. The BRG1 bromodomain exhibited binding, albeit weak, to acetylated peptides that were derived from histones H3 and H4. We have compared the acetyl-lysine binding sites of BRG1 bromodomain with the yGCN5 (general control of amino acid biosynthesis). By modeling the acetylated-lysine peptide into the BRG1 bromodomain structure, we were able to explain the weak binding of acetylated-lysine peptides to this bromodomain.
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http://dx.doi.org/10.1002/cbic.200600562DOI Listing
July 2007

Monitoring the effects of antagonists on protein-protein interactions with NMR spectroscopy.

J Am Chem Soc 2005 Sep;127(38):13220-6

Max Planck Institute for Biochemistry, D-82152 Martinsried, Germany.

We describe an NMR method that directly monitors the influence of ligands on protein-protein interactions. For a two-protein interaction complex, the size of one component should be small enough (less than ca. 15 kDa) to provide a good quality (15)N((13)C) HSQC spectrum after (15)N((13)C) labeling. The size of the second unlabeled component should be large enough so that the molecular weight of the preformed complex is larger than ca. 40 kDa. When the smaller protein binds to a larger one, broadening of NMR resonances results in the disappearance of most of its cross-peaks in the HSQC spectrum. Addition of an antagonist that can dissociate the complex would restore the HSQC spectrum of the smaller component. The method directly shows whether an antagonist releases proteins in their wild-type folded states or whether it induces their denaturation, partial unfolding, or precipitation. We illustrate the method by studying lead compounds that have recently been reported to block the MDM2-p53 interaction. Activation of p53 in tumor cells by inhibiting its interaction with MDM2 offers new strategy for cancer therapy.
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http://dx.doi.org/10.1021/ja052143xDOI Listing
September 2005

Molecular determinants for the complex formation between the retinoblastoma protein and LXCXE sequences.

J Biol Chem 2005 Nov 23;280(45):37868-76. Epub 2005 Aug 23.

Max Planck Institute for Biochemistry, Martinsried, Germany.

The retinoblastoma tumor suppressor protein (pRb) is a key negative regulator of cell proliferation that is frequently disregulated in human cancer. Many viral oncoproteins (for example, HPV E7 and E1A) are known to bind to the pRb pocket domain via a LXCXE binding motif. There are also some 20 cellular proteins that contain a LXCXE motif and have been reported to associate with the pocket domain of pRb. Using NMR spectroscopy and isothermal calorimetry titration, we show that LXCXE peptides of viral oncoproteins bind strongly to the pocket domain of pRb. Additionally, we show that LXCXE-like peptides of HDAC1 bind to the same site on pRb with a weak (micromolar) and transient association. Systematic substitution of residues other than conserved Leu, Cys, and Glu show that the residues flanking the LXCXE are important for the binding, whereas positively charged amino acids in the XLXCXEXXX sequence significantly weaken the interaction.
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http://dx.doi.org/10.1074/jbc.M504877200DOI Listing
November 2005

NMR and mass spectrometry studies of putative interactions of cell cycle proteins pRb and CDK6 with cell differentiation proteins MyoD and ID-2.

Biochim Biophys Acta 2005 Jun 14;1750(1):48-60. Epub 2005 Apr 14.

Max Planck Institute for Biochemistry, 82152 Martinsried, Germany.

Cell growth and differentiation require precise coordination of cell cycle and differentiation proteins. This can be achieved by direct interactions between proteins, by indirect interaction in multiprotein complexes, or by modulation of gene expression levels of partner proteins. Contradictory data abound in the literature regarding the binding between some central cell cycle proteins, pRb, and CDK6, with myogenic differentiation promoting, MyoD, and inhibiting, Id-2, factors. We have tested these interactions using pure proteins and in vitro biophysical and biochemical methods, which included mass spectrometry, nuclear magnetic resonance (NMR), the affinity chromatography pull-down assays, and gel filtration chromatography. Using this multimethod approach, we were able to document interactions between pRb and HPV-E7, pRb and SV40 large T antigen, CDK6 and p19, and MyoD and DNA. Using the same methods, we could unambiguously show that there is no direct protein-protein interaction in vitro between the small pocket domain of pRb and the bHLH domain of MyoD, the small pocket domain of pRb and Id-2, and CDK6 and a 15-amino-acid peptide from the C-terminal domain of MyoD. Indirect interactions, through additional binding partners in multiprotein complexes or modulation of gene expression levels of these proteins, are therefore their probable mode of action.
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http://dx.doi.org/10.1016/j.bbapap.2005.03.012DOI Listing
June 2005

NMR structural characterization of the N-terminal domain of the adenylyl cyclase-associated protein (CAP) from Dictyostelium discoideum.

J Biomol NMR 2004 May;29(1):73-84

Max Planck Institute for Biochemistry, 82152 Martinsried, Germany.

Cyclase-associated proteins (CAPs) are highly conserved, ubiquitous actin binding proteins that are involved in microfilament reorganization. The N-termini of CAPs play a role in Ras signaling and bind adenylyl cyclase; the C-termini bind to G-actin. We report here the NMR characterization of the amino-terminal domain of CAP from Dictyostelium discoideum (CAP(1-226)). NMR data, including the steady state (1)H-(15)N heteronuclear NOE experiments, indicate that the first 50 N-terminal residues are unstructured and that this highly flexible serine-rich fragment is followed by a stable, folded core starting at Ser 51. The NMR structure of the folded core is an alpha-helix bundle composed of six antiparallel helices, in a stark contrast to the recently determined CAP C-terminal domain structure, which is solely built by beta-strands.
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http://dx.doi.org/10.1023/B:JNMR.0000019513.86120.98DOI Listing
May 2004

1H, 15N and 13C NMR resonance assignments of staphostatin A, a specific Staphylococcus aureus cysteine proteinase inhibitor.

J Biomol NMR 2004 Mar;28(3):295-6

The increasing antibiotic resistance of an important human pathogen Staphylococcus aureus calls for the development of new therapeutic strategies. Staphylococcal cysteine proteases have been suggested as targets for such therapies. The recent discovery of staphostatins, specific protein inhibitors of these enzymes, gives prospects for the design and production of synthetic, low molecular weight analogs which might become drugs. We have decided to structurally characterize staphostatin A, a representative inhibitor of staphylococcal cysteine proteases, and to assess its binding mode to the target protease with the view of clarifying the specificity determinants. Here we report the (1)H, (15)N and (13)C NMR resonance assignments of staphostatin A.
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http://dx.doi.org/10.1023/B:JNMR.0000013681.37659.24DOI Listing
March 2004

A novel class of cysteine protease inhibitors: solution structure of staphostatin A from Staphylococcus aureus.

Biochemistry 2003 Nov;42(46):13449-56

Faculty of Biotechnology, Jagiellonian University, ul. Gronostajowa 7, 30-387 Krakow, Poland.

A series of secreted proteases are included among the virulence factors documented for Staphylococcus aureus. In light of increasing antibiotic resistance of this dangerous human pathogen, these proteases are considered as suitable targets for the development of novel therapeutic strategies. The recent discovery of staphostatins, endogenous, highly specific, staphylococcal cysteine protease inhibitors, opened a possibility for structure-based design of low molecular weight analogues. Moreover, the crystal structure of staphostatin B revealed a distinct folding pattern and an unexpected, substrate-like binding mode. The solution structure of staphostatin A reported here confirms that staphostatins constitute a novel, distinct class of cysteine protease inhibitors. In addition, the structure knowledge-based mutagenesis studies shed light on individual structural features of staphostatin A, the inhibition mechanism, and the determinants of distinct specificity of staphostatins toward their target proteases.
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http://dx.doi.org/10.1021/bi035310jDOI Listing
November 2003