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1,097 results on '"Schepartz, Alanna"'

202. Chemsearch: collaborative compound libraries with structure-aware browsing.

Author

Gaffney, Stephen G., Smaga, Sarah, Schepartz, Alanna, and Townsend, Jeffrey P.

Subjects
CHEMICALS, NAVIGATION, COMPUTER files, DIPLOIDY, ORGANIC compounds
Abstract

Summary: Chemsearch is a cross-platform server application for developing and managing a chemical compound library and associated data files, with an interface for browsing and search that allows for easy navigation to a com- pound of interest, similar compounds or compounds that have desired structural properties. With provisions for access control and centralized document and data storage, Chemsearch supports collaboration by distributed teams. [ABSTRACT FROM AUTHOR]

Published
2021
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204. Arginine topology controls escape of miniature proteins from early endosomes to the cytoplasm

Author

Appelbaum, Jacob S., LaRochelle, Jonathan R., Smith, Betsy A., Balkin, Daniel M., Holub, Justin M., and Schepartz, Alanna

Subjects
DNA-Binding Proteins, Cytoplasm, Protein Transport, Cations, Humans, Proteins, Zinc Fingers, Endosomes, Arginine, Article, HeLa Cells
Abstract

Proteins represent an expanding class of therapeutics, but their actions are limited primarily to extracellular targets because most peptidic molecules fail to enter cells. Here we identified two small proteins, miniature protein 5.3 and zinc finger module ZF5.3, that enter cells to reach the cytosol through rapid internalization and escape from Rab5+ endosomes. The trafficking pathway mapped for these molecules differs from that of Tat and Arg(8), which require transport beyond Rab5+ endosomes to gain cytosolic access. Our results suggest that the ability of 5.3 and ZF5.3 to escape from early endosomes is a unique feature and imply the existence of distinct signals, encodable within short sequences, that favor early versus late endosomal release. Identifying these signals and understanding their mechanistic basis will illustrate how cells control the movement of endocytic cargo and may allow researchers to engineer molecules to follow a desired delivery pathway for rapid cytosolic access.

Published
2012

205. A β-Boronopeptide Bundle of Known Structure As a Vehicle for Polyol Recognition

Author

Melicher, Michael S., Chu, John, Walker, Allison S., Miller, Scott J., Baxter, Richard H. G., and Schepartz, Alanna

Abstract

Despite significant progress in the design of receptors and sensors for simple polyols and monosaccharides, few synthetic receptors discriminate among multiple saccharide units simultaneously, especially under physiological conditions. Described here is the three-dimensional structure of a supramolecular complexa β-peptide bundledesigned for the potential to interact simultaneously with as many as eight discrete monosaccharide units. The preliminary evaluation of this construct as a vehicle for polyol binding is also presented.

Published
2024
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206. HOPS-Dependent Endosomal Escape Demands Protein Unfolding

Author

Zoltek, Madeline, Vázquez Maldonado, Angel L., Zhang, Xizi, Dadina, Neville, Lesiak, Lauren, and Schepartz, Alanna

Abstract

The inefficient translocation of proteins across biological membranes limits their application as potential therapeutics and research tools. In many cases, the translocation of a protein involves two discrete steps: uptake into the endocytic pathway and endosomal escape. Certain charged or amphiphilic molecules can achieve high protein uptake, but few are capable of efficient endosomal escape. One exception to this rule is ZF5.3, a mini-protein that exploits elements of the natural endosomal maturation machinery to translocate across endosomal membranes. Although some ZF5.3–protein conjugates are delivered efficiently to the cytosol or nucleus, overall delivery efficiency varies widely for different cargoes with no obvious design rules. Here we show that delivery efficiency depends on the ability of the cargo to unfold. Using fluorescence correlation spectroscopy, a single-molecule technique that precisely measures intracytosolic protein concentration, we show that regardless of size and pI, low-Tmcargoes of ZF5.3 (including intrinsically disordered domains) bias endosomal escape toward a high-efficiency pathway that requires the homotypic fusion and protein sorting (HOPS) complex. Small protein domains are delivered with moderate efficiency through the same HOPS portal, even if the Tmis high. These findings imply a novel pathway out of endosomes that is exploited by ZF5.3 and provide clear guidance for the selection or design of optimally deliverable therapeutic cargo.

Published
2024
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207. Dose-Dependent Nuclear Delivery and Transcriptional Repression with a Cell-Penetrant MeCP2

Author

Zhang, Xizi, Cattoglio, Claudia, Zoltek, Madeline, Vetralla, Carlo, Mozumdar, Deepto, and Schepartz, Alanna

Abstract

The vast majority of biologic-based therapeutics operate within serum, on the cell surface, or within endocytic vesicles, in large part because proteins and nucleic acids fail to efficiently cross cell or endosomal membranes. The impact of biologic-based therapeutics would expand exponentially if proteins and nucleic acids could reliably evade endosomal degradation, escape endosomal vesicles, and remain functional. Using the cell-permeant mini-protein ZF5.3, here we report the efficient nuclear delivery of functional Methyl-CpG-binding-protein 2 (MeCP2), a transcriptional regulator whose mutation causes Rett syndrome (RTT). We report that ZF-tMeCP2, a conjugate of ZF5.3 and MeCP2(Δaa13–71, 313–484), binds DNA in a methylation-dependent manner in vitro, and reaches the nucleus of model cell lines intact to achieve an average concentration of 700 nM. When delivered to live cells, ZF-tMeCP2 engages the NCoR/SMRT corepressor complex, selectively represses transcription from methylated promoters, and colocalizes with heterochromatin in mouse primary cortical neurons. We also report that efficient nuclear delivery of ZF-tMeCP2 relies on an endosomal escape portal provided by HOPS-dependent endosomal fusion. The Tat conjugate of MeCP2 (Tat-tMeCP2), evaluated for comparison, is degraded within the nucleus, is not selective for methylated promoters, and trafficks in a HOPS-independent manner. These results support the feasibility of a HOPS-dependent portal for delivering functional macromolecules to the cell interior using the cell-penetrant mini-protein ZF5.3. Such a strategy could broaden the impact of multiple families of biologic-based therapeutics.

Published
2024
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208. A Translation-Independent Directed Evolution Strategy to Engineer Aminoacyl-tRNA Synthetases

Author

Soni, Chintan, Prywes, Noam, Hall, Matthew, Nair, Malavika A., Savage, David F., Schepartz, Alanna, and Chatterjee, Abhishek

Abstract

Using directed evolution, aminoacyl-tRNA synthetases (aaRSs) have been engineered to incorporate numerous noncanonical amino acids (ncAAs). Until now, the selection of such novel aaRS mutants has relied on the expression of a selectable reporter protein. However, such translation-dependent selections are incompatible with exotic monomers that are suboptimal substrates for the ribosome. A two-step solution is needed to overcome this limitation: (A) engineering an aaRS to charge the exotic monomer, without ribosomal translation; (B) subsequent engineering of the ribosome to accept the resulting acyl-tRNA for translation. Here, we report a platform for aaRS engineering that directly selects tRNA-acylation without ribosomal translation (START). In START, each distinct aaRS mutant is correlated to a cognate tRNA containing a unique sequence barcode. Acylation by an active aaRS mutant protects the corresponding barcode-containing tRNAs from oxidative treatment designed to damage the 3′-terminus of the uncharged tRNAs. Sequencing of these surviving barcode-containing tRNAs is then used to reveal the identity of the aaRS mutants that acylated the correlated tRNA sequences. The efficacy of START was demonstrated by identifying novel mutants of the Methanomethylophilus alvuspyrrolysyl-tRNA synthetase from a naïve library that enables incorporation of ncAAs into proteins in living cells.

Published
2024
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213. Encodable activators of Src family kinases

Author

Zellefrow, Crystal D., Griffiths, Jennifer S., Saha, Sarmistha, Hodges, Abby M., Goodman, Jessica L., Paulk, Joshiawa, Kritzer, Joshua A., and Schepartz, Alanna

Subjects
Proline -- Chemical properties, Proline -- Research, Protein kinases -- Chemical properties, Protein kinases -- Research, Protein research, Chemistry
Abstract

A set of encodable miniature proteins that recognize SH3 domains from distinct Src family kinases with high affinity, where two of them activate Hck kinase with potencies that rival HIV Nef, which activates Hck kinase in vivo are described. SH3 domains are found to regulate the activity of Src family kinases through interaction with an internal proline-rich region that locks the kinase into a catalytically repressed state.

Published
2006

214. In Silico Improvement of beta(3)-Peptide Inhibitors of p53 center dot hDM2 and p53 center dot hDMX

Author

Michel, Julien, Harker, Elizabeth A., Tirado-Rives, Julian, Jorgensen, William L., and Schepartz, Alanna

Subjects
Colloid and Surface Chemistry, Chemistry(all), Biochemistry, Catalysis
Abstract

There is great interest in molecules capable of inhibiting the interactions between p53 and its negative regulators hDM2 and hDMX, as these molecules have validated potential against cancers in which one or both oncoproteins are overexpressed. We reported previously that appropriately substituted beta(3)-peptides inhibit these interactions and, more recently, that minimally cationic beta(3)-peptides are sufficiently cell permeable to upregulate p53-dependent genes in live cells. These observations, coupled with the known stability of beta-peptides in a cellular environment, and the recently reported structures of hDM2 and hDMX, motivated us to exploit computational modeling to identify beta-peptides with improved potency and/or selectivity. This exercise successfully identified a new beta(3)-peptide, beta 53-16, that possesses the highly desirable attribute of high affinity for both hDM2 and hDMX and identifies the 3,4-dichlorophenyl moiety as a novel determinant of hDMX affinity.

Published
2009
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215. Inhibiting HIV fusion with a beta-peptide foldamer

Author

Stephens, Olen M., Sunghwan, KIm, Hodsdon, Michael E., Welch, Brett D., Kay, Michael S., and Schepartz, Alanna

Subjects
Chemical synthesis -- Research, Aqueous solution reactions -- Research, Peptides -- Research, Chemistry
Abstract

A set of four beta(super 3)-peptides (betaWWW-1-4) is synthesized containing WW1epitope in both possible orientations on each available face of a beta(super 3)-decapeptide possessing significant 14-helix stability in aqueous solution due to electrostatic macrodipole stabilization and side chain-side chain salt bridges. The results show that molecules such as beta(super 3) peptides betaWW-1-4 could represent leads towards inhibitors or antigens effective against HIV or other viruses, such as SARS(super 30), Ebola, HRSV, and influenza that employ common fusion mechanisms.

Published
2005

216. Helical beta-peptide inhibitors of the p53-hDM2 interaction

Author

Kritzer, Joshua, Lear, James D., Hodson, Michael E., and Schepartz, Alanna

Subjects
Circular dichroism -- Research, Peptides -- Research, Peptides -- Structure, Chemistry
Abstract

Beta peptides differ from alpha peptides by one additional backbone carbon atom and by resistance to metabolism and proteolysis. A set of beta (super, 3) peptides that possess significant 14-helix structure in water that is one recognizes a cleft on the surface of the human oncogene product minute 2 (hDM2) with nanomolar affinity is reported.

Published
2004

217. Helix macrodipole control of beta(super 3)-peptide 14-helix stability in water

Author

Hart, Scott A., Bahadoor, Adilah B.F., Matthews, Erin E., Xiaoyan Qiu, and Schepartz, Alanna

Subjects
Carbonyl compounds -- Chemical properties, Water -- Composition, Peptides -- Research, Chemistry
Abstract

Beta-peptides attracted considerable attention by virtue of their ability to populate helical secondary structures, even in the absence of stabilizing tertiary interactions. The stabilization of the 14-helix macrodipole alleviates the requirement for multiple beta(super 3)-Glubeta(super 3)-Lys or beta(super 3)-Glu/beta(super 3)-Orn salt bridges on two of three helical faces is presented.

Published
2003

218. Confronting Racism in Chemistry Journals

Author

Burrows, Cynthia J., Huang, Jiaxing, Wang, Shu, Kim, Hyun Jae, Meyer, Gerald J., Schanze, Kirk, Lee, T. Randall, Lutkenhaus, Jodie L., Kaplan, David, Jones, Christopher, Bertozzi, Carolyn, Kiessling, Laura, Mulcahy, Mary Beth, Lindsley, Craig W., Finn, M. G., Blum, Joel D., Kamat, Prashant, Choi, Wonyong, Snyder, Shane, Aldrich, Courtney C., Rowan, Stuart, Liu, Bin, Liotta, Dennis, Weiss, Paul S., Zhang, Deqing, Ganesh, Krishna N., Atwater, Harry A., Gooding, J. Justin, Allen, David T., Voigt, Christopher A., Sweedler, Jonathan, Schepartz, Alanna, Rotello, Vincent, Lecommandoux, Sébastien, Sturla, Shana J., Hammes-Schiffer, Sharon, Buriak, Jillian, Steed, Jonathan W., Wu, Hongwei, Zimmerman, Julie, Brooks, Bryan, Savage, Phillip, Tolman, William, Hofmann, Thomas F., Brennecke, Joan F., Holme, Thomas A., Merz, Kenneth M., Scuseria, Gustavo, Jorgensen, William, Georg, Gunda I., Wang, Shaomeng, Proteau, Philip, Yates, John R., Stang, Peter, Walker, Gilbert C., Hillmyer, Marc, Taylor, Lynne S., Odom, Teri W., Carreira, Erick, Rossen, Kai, Chirik, Paul, Miller, Scott J., Shea, Joan-Emma, McCoy, Anne, Zanni, Martin, Hartland, Gregory, Scholes, Gregory, Loo, Joseph A., Milne, James, Tegen, Sarah B., Kulp, Daniel T., and Laskin, Julia

Published
2020
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219. Inhibiting epidermal growth factor receptor at a distance.

Author

Sinclair, Julie, Sinclair, Julie, Denton, Elizabeth, Schepartz, Alanna, Sinclair, Julie, Sinclair, Julie, Denton, Elizabeth, and Schepartz, Alanna

Abstract

The epidermal growth factor receptor (EGFR) tyrosine kinase is implicated in a large number of human cancers. Most EGFR inhibitors target the extracellular, growth factor-binding domain or the intracellular, ATP-binding domain. Here we describe molecules that inhibit the kinase activity of EGFR in a new way, by competing with formation of an essential intradimer coiled coil containing the juxtamembrane segment from each member of the receptor partnership. The most potent molecules we describe bind EGFR directly, decrease the proliferation of wild-type and mutant EGFR-dependent cells lines, inhibit phosphorylation of EGFR and downstream targets, and block coiled coil formation as judged by bipartite tetracysteine display. Potency is directly correlated with the ability to block coiled coil formation within full-length EGFR in cells.

Published
2014

220. Influence of macrocyclization on allosteric, juxtamembrane-derived, stapled peptide inhibitors of the epidermal growth factor receptor (EGFR).

Author

Sinclair, Julie K-L, Sinclair, Julie K-L, Schepartz, Alanna, Sinclair, Julie K-L, Sinclair, Julie K-L, and Schepartz, Alanna

Abstract

The hydrocarbon-stapled peptide E1(S) allosterically inhibits the kinase activity of the epidermal growth factor receptor (EGFR) by blocking a distant but essential protein-protein interaction: a coiled coil formed from the juxtamembrane segment (JM) of each member of the dimeric partnership.1 Macrocyclization is not required for activity: the analogous unstapled (but alkene-bearing) peptide is equipotent in cell viability, immunoblot, and bipartite display experiments to detect coiled coil formation on the cell surface.

Published
2014

221. Interactions of AsCy3 with cysteine-rich peptides.

Author

Alexander, Seth C, Alexander, Seth C, Schepartz, Alanna, Alexander, Seth C, Alexander, Seth C, and Schepartz, Alanna

Abstract

There is great interest in fluorogenic compounds that tag biomolecules within cells. Biarsenicals are fluorogenic compounds that become fluorescent upon binding four proximal Cys thiols, a tetracysteine (Cys(4)) motif. This work details interactions between the biarsenical AsCy3 and Cys(4) peptides. Maximal affinity was observed when two Cys-Cys pairs were separated by at least 8 amino acids; the highest affinity ligand bound in the nanomolar concentration range (K(app) = 43 nM) and with a significant (3.2-fold) fluorescence enhancement.

Published
2014

225. Highly specific DNA recognition by a designed miniature protein

Author

Zondlo, Neal J. and Schepartz, Alanna

Subjects
Immune recognition -- Research, DNA binding proteins -- Research, Chemistry
Abstract

Research was conducted to examine a novel strategy for the design of miniature proteins that present a solvent-exposed alpha-helix able to recognize DNA with high affinity and specificity. The approach involves dissecting those alpha-helical residues necessary for DNA recognition from their native protein context and grafting them on a stable, miniature protein core. Results indicate that the method may represent a general approach to the design of small, folded proteins that recognize nucleic acid and protein targets with high affinity and specificity.

Published
1999

227. Triplex tethered oligonucleotide probes

Author

Moses, Arikha C. and Schepartz, Alanna

Subjects
Nucleic acid probes -- Research, RNA -- Synthesis, Chemistry
Abstract

Tethered oligonucleotide probes (TOPs) can simultaneously recognize single- and double-stranded regions within a large, structured RNA in a Watson-Crick and Hoogsteen sense. These triplex TOPs form high affinity RNA complexes, with stabilities that are less sensitive to changes in temperature and monovalent cation concentration than model triple helices. Their ability to hybridize to single- and double-stranded regions of a target RNA indicates that they have potential use in the functional manipulation of RNAs in vitro and, with appropriate backbone modifications, in vivo.

Published
1996

232. Remodeling a β-peptide bundle

Author

Molski, Matthew A., primary, Goodman, Jessica L., additional, Chou, Fang-Chieh, additional, Baker, David, additional, Das, Rhiju, additional, and Schepartz, Alanna, additional

Published
2013
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