Protein domain mimetics as in vivo modulators of hypoxia-inducible factor signaling

Kushal, Swati; Lao, Brooke Bullock; Henchey, Laura K.; Dubey, Ramin; Mesallati, Hanah; Traaseth, Nathaniel J.; Olenyuk, Bogdan; Arora, Paramjit S.

doi:10.1073/pnas.1312473110

Cited by 92 publications

(141 citation statements)

References 47 publications

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“…Tryptophan fluorescence spectroscopy provides a dissociation constant, K d , of (3.8 ± 1.4) × 10 −8 M for HIF1α CTAD 786-826 to p300 CH1, which is consistent with the values obtained from a fluorescence polarization assay using fluorescein-labeled HIF1α CTAD (SI Appendix, Fig. S2) and those reported in the literature with isothermal titration microcalorimetry (14,19). OHM 1 targets CH1 with an affinity of (5.3 ± 1.4) × 10 −7 M ( Fig.…”

Section: Design and Synthesis Of Topographical Hif1αsupporting

confidence: 87%

“…1A). Small molecules that mimic the structural arrangement of the key residues on these helices should afford competitive inhibitors of HIF1α/p300 complex formation (19,22). We used a recently described strategy from our groups to mimic the interacting face of an α-helix on a small-molecule oxopiperazine scaffold (31).…”

Section: Design and Synthesis Of Topographical Hif1αmentioning

confidence: 99%

“…The binding affinities of OHMs for the p300-CH1 domain were evaluated using intrinsic tryptophan fluorescence spectroscopy, as described previously (19,34). Because Trp403 is located in the binding cleft of p300/CBP where a native HIF1α 816-824 helix binds, it offers a probe for investigating mimetics of this helix.…”

Section: Design and Synthesis Of Topographical Hif1αmentioning

confidence: 99%

“…Inhibitors of hypoxia-inducible gene expression would serve as unique tools for dissecting hypoxia signaling in tumors, as well as leads for cancer therapeutics (16)(17)(18)(19)(20)(21)(22)(23)(24)(25). The transcription factorcoactivator interaction presents an intriguing target for controlling hypoxia signaling because it is a critical node directing downstream expression of various genes that work in concert to modulate cancer progression.…”

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confidence: 99%

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In vivo modulation of hypoxia-inducible signaling by topographical helix mimetics

Lao

Grishagin

Mesallati

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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126

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Development of small-molecule inhibitors of protein-protein interactions is a fundamental challenge at the interface of chemistry and cancer biology. Successful methods for design of protein-protein interaction inhibitors include computational and experimental high-throughput and fragment-based screening strategies to locate small-molecule fragments that bind protein surfaces. An alternative rational design approach seeks to mimic the orientation and disposition of critical binding residues at protein interfaces. We describe the design, synthesis, biochemical, and in vivo evaluation of a small-molecule scaffold that captures the topography of α-helices. We designed mimics of a key α-helical domain at the interface of hypoxia-inducible factor 1α and p300 to develop inhibitors of hypoxia-inducible signaling. The hypoxia-inducible factor/ p300 interaction regulates the transcription of key genes, whose expression contributes to angiogenesis, metastasis, and altered energy metabolism in cancer. The designed compounds target the desired protein with high affinity and in a predetermined manner, with the optimal ligand providing effective reduction of tumor burden in experimental animal models.helix mimics | hypoxia signaling | synthetic inhibitors of transcription

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Section: Design and Synthesis Of Topographical Hif1αsupporting

confidence: 87%

Section: Design and Synthesis Of Topographical Hif1αmentioning

confidence: 99%

Section: Design and Synthesis Of Topographical Hif1αmentioning

confidence: 99%

mentioning

confidence: 99%

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In vivo modulation of hypoxia-inducible signaling by topographical helix mimetics

Lao

Grishagin

Mesallati

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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126

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“…of these synthetic helices by CD and 2D-NMR spectroscopy, as well as single crystal X-ray diffraction analysis, reveals that they faithfully mimic the conformation of canonical α-helices (21,22). Importantly, HBS helices are able to inhibit intracellular protein-protein interactions mediated by α-helical domains, supporting the hypothesis that these compounds reproduce the structure and function of protein α-helices (23,24). To analyze the effect of different residues on α-helix formation, we prepared an HBS analog with a disulfide linkage whose rates of formation could be monitored under aqueous conditions (25).…”

Section: Significancementioning

confidence: 67%

Effects of side chains in helix nucleation differ from helix propagation

Miller

Watkins

Kallenbach

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Helix-coil transition theory connects observable properties of the α-helix to an ensemble of microstates and provides a foundation for analyzing secondary structure formation in proteins. Classical models account for cooperative helix formation in terms of an energetically demanding nucleation event (described by the σ constant) followed by a more facile propagation reaction, with corresponding s constants that are sequence dependent. Extensive studies of folding and unfolding in model peptides have led to the determination of the propagation constants for amino acids. However, the role of individual side chains in helix nucleation has not been separately accessible, so the σ constant is treated as independent of sequence. We describe here a synthetic model that allows the assessment of the role of individual amino acids in helix nucleation. Studies with this model lead to the surprising conclusion that widely accepted scales of helical propensity are not predictive of helix nucleation. Residues known to be helix stabilizers or breakers in propagation have only a tenuous relationship to residues that favor or disfavor helix nucleation.synthetic helices | helix propensity T he α-helix is the most prevalent secondary structure in proteins and can form extremely rapidly. Helix formation is thus crucial in early steps of protein folding, and a complete description of the kinetics and thermodynamics of α-helix formation is fundamental for understanding protein folding (1). Theoretical models of the helix-coil transition consider helix formation to proceed in two steps: initial nucleation of a helical sequence, denoted by the parameter σ in the Zimm-Bragg notation (2), followed by more favorable helix propagation, denoted by s parameters. This distinction identifies the energetically unfavorable organization of three consecutive amino acid residues to form a helical nucleus as the slow step in helix formation (2-4). Helix propagation, in contrast, refers to the addition of the next hydrogen bond to a preformed helix (Fig. 1A). Zimm-Bragg or equivalent theories posit that formation of short peptide helices in water is unfavorable because the large decrease in entropy required for nucleation is not adequately compensated by the enthalpic gain from forming a small number of hydrogen bonds.The ability of different peptide sequences to adopt helical conformations has been rigorously investigated, and the stability of α-helices can be estimated from widely used scales of helical propensity (5). These scales are important for our understanding of protein structure, folding, and function. The classical studies for determination of helix propensities have used peptides, coiled-coils, and protein models for host-guest studies in which a guest residue is systematically substituted at a site in a host structure (6-12). Helix-coil transition theory then relates changes in the conformational stability to microscopic helix propensities or s constants for individual residues. The results provide a quantitative basis for evaluating t...

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Synthesis of Hydrogen‐Bond Surrogate α‐Helices as Inhibitors of Protein‐Protein Interactions

Miller

Thomson

Arora

2014

CP Chemical Biology

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The α‐helix is a prevalent secondary structure in proteins and is critical in mediating protein‐protein interactions (PPIs). Peptide mimetics that adopt stable helices have become powerful tools for the modulation of PPIs in vitro and in vivo. Hydrogen‐bond surrogate (HBS) α‐helices utilize a covalent bond in place of an N‐terminal i to i+4 hydrogen bond and have been used to target and disrupt PPIs that become dysregulated in disease states. These compounds have improved conformational stability and cellular uptake as compared to their linear peptide counterparts. The protocol presented here describes current methodology for the synthesis of HBS α‐helical mimetics. The solid‐phase synthesis of HBS helices involves solid‐phase peptide synthesis with three key steps involving incorporation of N‐allyl functionality within the backbone of the peptide, coupling of a secondary amine, and a ring‐closing metathesis step. Curr. Protoc. Chem. Biol. 6:101‐116 © 2014 by John Wiley & Sons, Inc.

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Protein domain mimetics as in vivo modulators of hypoxia-inducible factor signaling

Cited by 92 publications

References 47 publications

In vivo modulation of hypoxia-inducible signaling by topographical helix mimetics

In vivo modulation of hypoxia-inducible signaling by topographical helix mimetics

Effects of side chains in helix nucleation differ from helix propagation

Synthesis of Hydrogen‐Bond Surrogate α‐Helices as Inhibitors of Protein‐Protein Interactions

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