TY - JOUR A1 - Zhang, Fuzhong Z. A1 - Timm, Katharina A. A1 - Arndt, Katja Maren A1 - Woolley, G. Andrew T1 - Photocontrol of Coiled-Coil Proteins in Living Cells N2 - Light switching of the activity of a coiled-coil protein, the AP-1 transcription factor, in living cells was made possible by the introduction of a designed azobenzene-cross-linked dominant negative peptide, XAFosW (red and yellow in the picture). In the dark, XAFosW showed decreased helical content and decreased affinity for target Jun proteins (green); irradiation at 365 nm enhanced helicity and target affinity. Y1 - 2010 UR - http://www3.interscience.wiley.com/cgi-bin/jhome/26737/ U6 - https://doi.org/10.1002/anie.201000909 SN - 1433-7851 ER - TY - CHAP A1 - Rejiba, Soukaina A1 - Frelan, Megan A1 - Hermann, Alex A1 - Arndt, Katja Maren A1 - Hajri, Amor T1 - Interfering peptides targeting transcription factor AP1 for pancreatic cancer gene therapy T2 - Molecular therapy : the journal of the American Society of Gene Therapy Y1 - 2013 SN - 1525-0016 VL - 21 IS - 2 SP - S250 EP - S250 PB - Nature Publ. Group CY - New York ER - TY - JOUR A1 - Speck, Janina A1 - Arndt, Katja Maren A1 - Müller, Kristian M. T1 - Efficient phage display of intracellularly folded proteins mediated by the TAT pathway JF - Protein engineering design & selection N2 - Phage display with filamentous phages is widely applied and well developed, yet proteins requiring a cytoplasmic environment for correct folding still defy attempts at functional display. To extend applicability of phage display, we employed the twin-arginine translocation (TAT) pathway to incorporate proteins fused to the C-terminal domain of the geneIII protein into phage particles. We investigated functionality and display level of fluorescent proteins depending on the translocation pathway, which was the TAT, general secretory (SEC) or signal recognition particle (SRP) pathway mediated by the TorA, PelB or DsbA signal sequences, respectively. Importantly, for green fluorescent protein, yellow fluorescent protein and cyan fluorescent protein, only TAT, but not SEC or SRP, translocation led to fluorescence of purified phage particles, although all three proteins could be displayed regardless of the translocation pathway. In contrast, the monomeric red fluorescent protein mCherry was functionally displayed regardless of the translocation pathway. Hence, correct folding and fluorophor formation of mCherry is not limited to the cytosol. Furthermore, we successfully displayed firefly luciferase as well as an 83 kDa argonaute protein, both containing free cysteines. This demonstrates broad applicability of the TAT-mediated phagemid system for the display of proteins requiring cytoplasmic factors for correct folding and should prove useful for the display of proteins requiring incorporation of co-factors or oligomerization to gain function. KW - g3p KW - phagemid display KW - protein design KW - protein engineering KW - selection Y1 - 2011 U6 - https://doi.org/10.1093/protein/gzr001 SN - 1741-0126 VL - 24 IS - 6 SP - 473 EP - 484 PB - Oxford Univ. Press CY - Oxford ER - TY - CHAP A1 - Kükenshöner, Tim A1 - Jean-Christoph, N. A1 - Speck, J. A1 - Müller, Kristian M. A1 - Arndt, Katja Maren T1 - Targeting the microphthalmia associated transcription factor coiled coil domain with interfering peptides T2 - The FEBS journal Y1 - 2011 SN - 1742-464X VL - 278 IS - 6 SP - 159 EP - 159 PB - Wiley-Blackwell CY - Malden ER - TY - JOUR A1 - Azuma, Yusuke A1 - Kuekenshoener, Tim A1 - Ma, Guangyong A1 - Yasunaga, Jun-ichiro A1 - Imanishi, Miki A1 - Tanaka, Gen A1 - Nakase, Ikuhiko A1 - Maruno, Takahiro A1 - Kobayashi, Yuji A1 - Arndt, Katja Maren A1 - Matsuoka, Masao A1 - Futaki, Shiroh T1 - Controlling leucine-zipper partner recognition in cells through modification of a-g interactions JF - Chemical communications N2 - By focusing on the a-g interactions, successful design and selection were accomplished to obtain a leucine-zipper segment that discriminates the appropriate partner over another that provides very similar patterns of electrostatic interactions. Y1 - 2014 U6 - https://doi.org/10.1039/c4cc00555d SN - 1359-7345 SN - 1364-548X VL - 50 IS - 48 SP - 6364 EP - 6367 PB - Royal Society of Chemistry CY - Cambridge ER - TY - JOUR A1 - Hagen, Sven A1 - Baumann, Tobias A1 - Wagner, Hanna J. A1 - Morath, Volker A1 - Kaufmann, Beate A1 - Fischer, Adrian A1 - Bergmann, Stefan A1 - Schindler, Patrick A1 - Arndt, Katja Maren A1 - Mueller, Kristian M. T1 - Modular adeno-associated virus (rAAV) vectors used for cellular virus-directed enzyme prodrug therapy JF - Scientific reports N2 - The pre-clinical and clinical development of viral vehicles for gene transfer increased in recent years, and a recombinant adeno-associated virus (rAAV) drug took center stage upon approval in the European Union. However, lack of standardization, inefficient purification methods and complicated retargeting limit general usability. We address these obstacles by fusing rAAV-2 capsids with two modular targeting molecules (DARPin or Affibody) specific for a cancer cell-surface marker (EGFR) while simultaneously including an affinity tag (His-tag) in a surface-exposed loop. Equipping these particles with genes coding for prodrug converting enzymes (thymidine kinase or cytosine deaminase) we demonstrate tumor marker specific transduction and prodrug-dependent apoptosis of cancer cells. Coding terminal and loop modifications in one gene enabled specific and scalable purification. Our genetic parts for viral production adhere to a standardized cloning strategy facilitating rapid prototyping of virus directed enzyme prodrug therapy (VDEPT). Y1 - 2014 U6 - https://doi.org/10.1038/srep03759 SN - 2045-2322 VL - 4 PB - Nature Publ. Group CY - London ER - TY - JOUR A1 - Mazumder, Mostafizur A1 - Brechun, Katherine E. A1 - Kim, Yongjoo B. A1 - Hoffmann, Stefan A. A1 - Chen, Yih Yang A1 - Keiski, Carrie-Lynn A1 - Arndt, Katja Maren A1 - McMillen, David R. A1 - Woolley, G. Andrew T1 - An Escherichia coli system for evolving improved light-controlled DNA-binding proteins JF - Protein engineering design & selection N2 - Light-switchable proteins offer numerous opportunities as tools for manipulating biological systems with exceptional degrees of spatiotemporal control. Most designed light-switchable proteins currently in use have not been optimised using the randomisation and selection/screening approaches that are widely used in other areas of protein engineering. Here we report an approach for screening light-switchable DNA-binding proteins that relies on light-dependent repression of the transcription of a fluorescent reporter. We demonstrate that the method can be used to recover a known light-switchable DNA-binding protein from a random library. KW - directed evolution KW - fluorescent reporter KW - optogenetics Y1 - 2015 U6 - https://doi.org/10.1093/protein/gzv033 SN - 1741-0126 SN - 1741-0134 VL - 28 IS - 9 SP - 293 EP - 302 PB - Oxford Univ. Press CY - Oxford ER - TY - JOUR A1 - Zuo, Zhili A1 - Gandhi, Neha S. A1 - Arndt, Katja Maren A1 - Mancera, Ricardo L. T1 - Free energy calculations of the interactions of c-Jun-based synthetic peptides with the c-Fos protein JF - Biopolymers N2 - The c-Fosc-Jun complex forms the activator protein 1 transcription factor, a therapeutic target in the treatment of cancer. Various synthetic peptides have been designed to try to selectively disrupt the interaction between c-Fos and c-Jun at its leucine zipper domain. To evaluate the binding affinity between these synthetic peptides and c-Fos, polarizable and nonpolarizable molecular dynamics (MD) simulations were conducted, and the resulting conformations were analyzed using the molecular mechanics generalized Born surface area (MM/GBSA) method to compute free energies of binding. In contrast to empirical and semiempirical approaches, the estimation of free energies of binding using a combination of MD simulations and the MM/GBSA approach takes into account dynamical properties such as conformational changes, as well as solvation effects and hydrophobic and hydrophilic interactions. The predicted binding affinities of the series of c-Jun-based peptides targeting the c-Fos peptide show good correlation with experimental melting temperatures. This provides the basis for the rational design of peptides based on internal, van der Waals, and electrostatic interactions. KW - free energy of binding KW - coiled-coil KW - molecular dynamics KW - MM KW - GBSA KW - leucine zipper Y1 - 2012 U6 - https://doi.org/10.1002/bip.22099 SN - 0006-3525 VL - 97 IS - 11 SP - 899 EP - 909 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Hagen, Sven A1 - Mattay, Dinah A1 - Raeuber, Christina A1 - Mueller, Kristian M. A1 - Arndt, Katja Maren T1 - Characterization and inhibition of AF10-mediated interaction JF - Journal of peptide science N2 - The non-random chromosomal translocations t(10;11)(p13;q23) and t(10;11)(p13;q14-21) result in leukemogenic fusion proteins comprising the coiled coil domain of the transcription factor AF10 and the proteins MLL or CALM, respectively, and subsequently cause certain types of acute leukemia. The AF10 coiled-coil domain, which is crucial for the leukemogenic effect, has been shown to interact with GAS41, a protein previously identified as the product of an amplified gene in glioblastoma. Using sequential synthetic peptides, we mapped the potential AF10/GAS41 interaction site, which was subsequently be used as scaffold for a library targeting the AF10 coiled-coil domain. Using phage display, we selected a peptide that binds the AF10 coiled-coil domain with higher affinity than the respective coiled-coil region of wild-type GAS41, as demonstrated by phage ELISA, CD, and PCAs. Furthermore, we were able to successfully deploy the inhibitory peptide in a mammalian cell line to lower the expression of Hoxa genes that have been described to be overexpressed in these leukemias. This work dissects molecular determinants mediating AF10-directed interactions in leukemic fusions comprising the N-terminal parts of the proteins MLL or CALM and the C-terminal coiled-coil domain of AF10. Furthermore, it outlines the first steps in recognizing and blocking the leukemia-associated AF10 interaction in histiocytic lymphoma cells and therefore, may have significant implications in future diagnostics and therapeutics. Copyright (c) 2014 European Peptide Society and John Wiley & Sons, Ltd. KW - protein-protein interaction KW - protein design and selection KW - protein engineering KW - coiled coil KW - leucine zipper KW - AF10 Y1 - 2014 U6 - https://doi.org/10.1002/psc.2626 SN - 1075-2617 SN - 1099-1387 VL - 20 IS - 6 SP - 385 EP - 397 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Kuekenshoener, Tim A1 - Wohlwend, Daniel A1 - Niemoeller, Christoph A1 - Dondapati, Padmarupa A1 - Speck, Janina A1 - Adeniran, Adebola V. A1 - Nieth, Anita A1 - Gerhardt, Stefan A1 - Einsle, Oliver A1 - Mueller, Kristian M. A1 - Arndt, Katja Maren T1 - Improving coiled coil stability while maintaining specificity by a bacterial hitchhiker selection system JF - Journal of structural biology N2 - The design and selection of peptides targeting cellular proteins is challenging and often yields candidates with undesired properties. Therefore we deployed a new selection system based on the twin-arginine translocase (TAT) pathway of Escherichia coli, named hitchhiker translocation (HiT) selection. A pool of alpha-helix encoding sequences was designed and selected for interference with the coiled coil domain (CC) of a melanoma-associated basic-helix-loop-helix-leucine-zipper (bHLHLZ) protein, the microphthalmia associated transcription factor (MITF). One predominant sequence (iM10) was enriched during selection and showed remarkable protease resistance, high solubility and thermal stability while maintaining its specificity. Furthermore, it exhibited nanomolar range affinity towards the target peptide. A mutation screen indicated that target-binding helices of increased homodimer stability and improved expression rates were preferred in the selection process. The crystal structure of the iM10/MITF-CC heterodimer (2.1 angstrom) provided important structural insights and validated our design predictions. Importantly, iM10 did not only bind to the MITF coiled coil, but also to the markedly more stable HLHLZ domain of MITF. Characterizing the selected variants of the semi-rational library demonstrated the potential of the innovative bacterial selection approach. (C) 2014 Elsevier Inc. All rights reserved. KW - Basic helix-loop-helix leucine zipper KW - Coiled coils KW - Microphthalmia associated transcription factor KW - Selection and design KW - Twin arginine translocation pathway Y1 - 2014 U6 - https://doi.org/10.1016/j.jsb.2014.03.002 SN - 1047-8477 SN - 1095-8657 VL - 186 IS - 3 SP - 335 EP - 348 PB - Elsevier CY - San Diego ER -