47 research outputs found
Semi-Quantitative Models for Identifying Potent and Selective Transthyretin Amyloidogenesis Inhibitors
Rate-limiting dissociation of the tetrameric protein transthyretin (TTR), followed by monomer misfolding and misassembly, appears to cause degenerative diseases in humans known as the transthyretin amyloidoses, based on human genetic, biochemical and pharmacologic evidence. Small molecules that bind to the generally unoccupied thyroxine binding pockets in the native TTR tetramer kinetically stabilize the tetramer, slowing subunit dissociation proportional to the extent that the molecules stabilize the native state over the dissociative transition state—thereby inhibiting amyloidogenesis. Herein, we use previously reported structure-activity relationship data to develop two semi-quantitative algorithms for identifying the structures of potent and selective transthyretin kinetic stabilizers/amyloidogenesis inhibitors. The viability of these prediction algorithms, in particular the more robust in silico docking model, is perhaps best validated by the clinical success of tafamidis, the first-in-class drug approved in Europe, Japan, South America, and elsewhere for treating transthyretin aggregation-associated familial amyloid polyneuropathy. Tafamidis is also being evaluated in a fully-enrolled placebo-controlled clinical trial for its efficacy against TTR cardiomyopathy. These prediction algorithms will be useful for identifying second generation TTR kinetic stabilizers, should these be needed to ameliorate the central nervous system or ophthalmologic pathology caused by TTR aggregation in organs not accessed by oral tafamidis administration
Biological Imaging Capability in the ABRS Facility on ISS
This slide presentation reviews the Advanced Biological Research System (ABRS) on the International Space Station (ISS) and its biological imaging capability. The ABRS is an environmental control chamber. It has two indpendently controlled Experiment Research Chambers (ERCs) with temperature, relative humidity and carbon dioxide controls. ABRS is a third generation plant growth system. Several experiments are reviewed, with particular interest in the use of Green Fluorescent Protein (GFP) a non-destructive plant stress reporting mechanism, naturally found in jellyfish
The genomic landscape of balanced cytogenetic abnormalities associated with human congenital anomalies
Despite the clinical significance of balanced chromosomal abnormalities (BCAs), their characterization has largely been restricted to cytogenetic resolution. We explored the landscape of BCAs at nucleotide resolution in 273 subjects with a spectrum of congenital anomalies. Whole-genome sequencing revised 93% of karyotypes and demonstrated complexity that was cryptic to karyotyping in 21% of BCAs, highlighting the limitations of conventional cytogenetic approaches. At least 33.9% of BCAs resulted in gene disruption that likely contributed to the developmental phenotype, 5.2% were associated with pathogenic genomic imbalances, and 7.3% disrupted topologically associated domains (TADs) encompassing known syndromic loci. Remarkably, BCA breakpoints in eight subjects altered a single TAD encompassing MEF2C, a known driver of 5q14.3 microdeletion syndrome, resulting in decreased MEF2C expression. We propose that sequence-level resolution dramatically improves prediction of clinical outcomes for balanced rearrangements and provides insight into new pathogenic mechanisms, such as altered regulation due to changes in chromosome topology
Effects of Single α‑to‑β Residue Replacements on Structure and Stability in a Small Protein: Insights from Quasiracemic Crystallization
Synthetic peptides
that contain backbone modifications but nevertheless adopt folded
structures similar to those of natural polypeptides are of fundamental
interest and may provide a basis for biomedical applications. Such
molecules can, for example, mimic the ability of natural prototypes
to bind to specific target macromolecules but resist degradation by
proteases. We have previously shown that oligomers containing mixtures
of α- and β-amino acid residues (“α/β-peptides”)
can mimic the α-helix secondary structure, and that properly
designed α/β-peptides can bind to proteins that evolved
to bind to α-helical partners. Here we report fundamental studies
that support the long-range goal of extending the α/β
approach to tertiary structures. We have evaluated the impact of single
α → β modifications on the structure and stability
of the small and well-studied villin headpiece subdomain (VHP). The
native state of this 35-residue polypeptide contains several α-helical
segments packed around a small hydrophobic core. We examined α
→ β substitution at four solvent-exposed positions, Asn19,
Trp23, Gln26 and Lys30. In each case, both the β<sup>3</sup> homologue of the natural α residue and a cyclic β residue
were evaluated. All α → β<sup>3</sup> substitutions
caused significant destabilization of the tertiary structure as measured
by variable-temperature circular dichroism, although at some of these
positions, replacing the β<sup>3</sup> residue with a cyclic
β residue led to improved stability. Atomic-resolution structures
of four VHP analogues were obtained via quasiracemic crystallization.
These findings contribute to a fundamental α/β-peptide
knowledge-base by confirming that β<sup>3</sup>-amino acid residues
can serve as effective structural mimics of homologous α-amino
acid residues within a natural tertiary fold, which should support
rational design of functional α/β analogues of natural
poly-α-peptides
Evidence for Phenylalanine Zipper-Mediated Dimerization in the X‑ray Crystal Structure of a Magainin 2 Analogue
High-resolution
structure elucidation has been challenging for
the large group of host-defense peptides that form helices on or within
membranes but do not manifest a strong folding propensity in aqueous
solution. Here we report the crystal structure of an analogue of the
widely studied host-defense peptide magainin 2. Magainin 2 (S8A, G13A,
G18A) is a designed variant that displays enhanced antibacterial activity
relative to the natural peptide. The crystal structure of magainin
2 (S8A, G13A, G18A), obtained for the racemic form, features a dimerization
mode that has previously been proposed to play a role in the antibacterial
activity of magainin 2 and related peptides
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Using sulfuramidimidoyl fluorides that undergo sulfur(VI) fluoride exchange for inverse drug discovery.
Drug candidates that form covalent linkages with their target proteins have been underexplored compared with the conventional counterparts that modulate biological function by reversibly binding to proteins, in part due to concerns about off-target reactivity. However, toxicity linked to off-target reactivity can be minimized by using latent electrophiles that only become activated towards covalent bond formation on binding a specific protein. Here we study sulfuramidimidoyl fluorides, a class of weak electrophiles that undergo sulfur(VI) fluoride exchange chemistry. We show that equilibrium binding of a sulfuramidimidoyl fluoride to a protein can allow nucleophilic attack by a specific amino acid side chain, which leads to conjugate formation. We incubated small molecules, each bearing a sulfuramidimidoyl fluoride electrophile, with human cell lysate, and the protein conjugates formed were identified by affinity chromatography-mass spectrometry. This inverse drug discovery approach identified a compound that covalently binds to and irreversibly inhibits the activity of poly(ADP-ribose) polymerase 1, an important anticancer target in living cells
Enhancement of α-Helix Mimicry by an α/β-Peptide Foldamer via Incorporation of a Dense Ionic Side-Chain Array
We report a new method for preorganization of α/β-peptide
helices, based on the use of a dense array of acidic and basic side
chains. Previously we have used cyclically constrained β residues
to promote α/β-peptide helicity; here we show that an
engineered ion pair array can be comparably effective, as indicated
by mimicry of the CHR domain of HIV protein gp41. The new design is
effective in biochemical and cell-based infectivity assays; however,
the resulting α/β-peptide is susceptible to proteolysis.
This susceptibility was addressed via introduction of a few cyclic
β residues near the cleavage site, to produce the most stable,
effective α/β-peptide gp41 CHR analogue identified. Crystal
structures of an α- and α/β-peptide (each involved
in a gp41-mimetic helix bundle) that contain the dense acid/base residue
array manifest disorder in the ionic side chains, but there is little
side-chain disorder in analogous α- and α/β-peptide
structures with a sparser ionic side-chain array. These observations
suggest that dense arrays of complementary acidic and basic residues
can provide conformational stabilization via Coulombic attractions
that do not require entropically costly ordering of side chains