51 research outputs found
Retinoid isomerase inhibitors impair but do not block mammalian cone photoreceptor function
Visual function in vertebrates critically depends on the continuous regeneration of visual pigments in rod and cone photoreceptors. RPE65 is a well-established retinoid isomerase in the pigment epithelium that regenerates rhodopsin during the rod visual cycle; however, its contribution to the regeneration of cone pigments remains obscure. In this study, we use potent and selective RPE65 inhibitors in rod- and cone-dominant animal models to discern the role of this enzyme in cone-mediated vision. We confirm that retinylamine and emixustat-family compounds selectively inhibit RPE65 over DES1, the putative retinoid isomerase of the intraretinal visual cycle. In vivo and ex vivo electroretinography experiments in Gnat1-/- mice demonstrate that acute administration of RPE65 inhibitors after a bleach suppresses the late, slow phase of cone dark adaptation without affecting the initial rapid portion, which reflects intraretinal visual cycle function. Acute administration of these compounds does not affect the light sensitivity of cone photoreceptors in mice during extended exposure to background light, but does slow all phases of subsequent dark recovery. We also show that cone function is only partially suppressed in cone-dominant ground squirrels and wild-type mice by multiday administration of an RPE65 inhibitor despite profound blockade of RPE65 activity. Complementary experiments in these animal models using the DES1 inhibitor fenretinide show more modest effects on cone recovery. Collectively, these studies demonstrate a role for continuous RPE65 activity in mammalian cone pigment regeneration and provide further evidence for RPE65-independent regeneration mechanisms
Ubistatins Inhibit Proteasome-Dependent Degradation by Binding the Ubiquitin Chain
To identify previously unknown small molecules that inhibit cell cycle machinery, we performed a chemical genetic screen in Xenopus extracts. One class of inhibitors, termed ubistatins, blocked cell cycle progression by inhibiting cyclin B proteolysis and inhibited degradation of ubiquitinated Sic1 by purified proteasomes. Ubistatins blocked the binding of ubiquitinated substrates to the proteasome by targeting the ubiquitin-ubiquitin interface of Lys^(48)-linked chains. The same interface is recognized by ubiquitin-chain receptors of the proteasome, indicating that ubistatins act by disrupting a critical protein-protein interaction in the ubiquitin-proteasome system
A small molecule agonist of EphA2 receptor tyrosine kinase inhibits tumor cell migration in vitro and prostate cancer metastasis in vivo
10.1371/journal.pone.0042120PLoS ONE78
New Methodology toward α,β-Unsaturated Carboxylic Acids from Saturated Acids
A carefully
designed three-step unsaturation of carboxylic acids
is described. Briefly, carboxylic acids were converted to the trifluoromethyl
ketone. Subsequent treatment with selenium dioxide followed by hydrolysis
afforded α,β-unsaturated carboxylic acids. The mechanism
of the reported transformation was investigated, which led us to propose
a novel explanation featuring selenium dioxide assisted enolizaion
of a trifluoromethyl ketone followed by β-deprotonation
Bile Acid Recognition by Mouse Ileal Bile Acid Binding Protein
Ileal bile acid binding
protein (I-BABP, gene name FABP6) is a component of the bile acid
recycling system, expressed in the ileal enterocyte. The physiological
role of I-BABP has been hypothesized to be either an intracellular
buffering agent to protect against excess intracellular bile acids
or separately as a modulator of bile acid controlled transcription.
We investigated mouse I-BABP (mI-BABP) to understand the function
of this protein family. Here, we studied energetics and site selectivity
of binding with physiological bile acids using a combination of isothermal
calorimetric analysis and NMR spectroscopy. We found that the most
abundant bile acid in the mouse (β-muricholic acid) binds with
weak affinity individually and in combination with other bile acids.
Further analysis showed that mI-BABP like human I-BABP (hI-BABP) specifically
recognizes the conjugated form of cholic acid:chenodeoxycholic acid
(CA:CDCA) in a site-selective manner, displaying the highest affinity
of any bile acid combination tested. These results indicate that I-BABP
specifically recognizes the ligand combination of CDCA and CA, even
in a species such as the mouse where CDCA only represents a trace
component of the physiological pool. Specific and conserved recognition
of the CDCA and CA ligand combination suggests that I-BABP may play
a critical role in the regulation of bile acid signaling in addition
to its proposed role as a buffering agent
Approach for Expanding Triterpenoid Complexity via Divergent Norrish-Yang Photocyclization
Triterpenoids comprise a very diverse
family of polycyclic molecules
that is well-known to possess a myriad of medicinal properties. Therefore,
triterpenoids constitute an attractive target for medicinal chemistry
and diversity-oriented synthesis. Photochemical transformations provide
a promising tool for the rapid, green, and inexpensive generation
of skeletal diversity in the construction of natural product-like
libraries. With this in mind, we have developed a diversity-oriented
strategy, whereby the parent triterpenoids bryonolic acid and lanosterol
are converted to the pseudosymmetrical polyketones by sequential allylic
oxidation and oxidative cleavage of the bridging double bond at the
B/C ring fusion. The resultant polyketones were hypothesized to undergo
divergent Norrish-Yang cyclization to produce unique 6/4/8-fused triterpenoid
analogues. The subtle differences between parent triterpenoids led
to dramatically different spatial arrangements of reactive functionalities.
This finding was rationalized through conformational analysis to explain
unanticipated photoinduced pinacolization, as well as the regio- and
stereochemical outcome of the desired Norrish-Yang cyclization
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Catalytic synthesis of 9- cis -retinoids: mechanistic insights
The regioselective Z-isomerization of thermodynamically stable all-trans retinoids remains challenging, and ultimately limits the availability of much needed therapeutics for the treatment of human diseases. We present here a novel, straightforward approach for the catalytic Z-isomerization of retinoids using conventional heat treatment or microwave irradiation. A screen of 20 transition metal-based catalysts identified an optimal approach for the regioselective production of Z-retinoids. The most effective catalytic system was comprised of a palladium complex with labile ligands. Several mechanistic studies, including isotopic H/D exchange and state-of-the-art quantum chemical calculations using coupled cluster methods indicate that the isomerization is initiated by catalyst dimerization followed by the formation of a cyclic, six-membered chloropalladate catalyst-substrate adduct, which eventually opens to produce the desired Z-isomer. The synthetic development described here, combined with thorough mechanistic analysis of the underlying chemistry, highlights the use of readily available transition metal-based catalysts in straightforward formats for gram-scale drug synthesis
A Single Hydroxyl Group Governs Ligand Site Selectivity in Human Ileal Bile Acid Binding Protein
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Z -isomerization of retinoids through combination of monochromatic photoisomerization and metal catalysis
Catalytic Z-isomerization of retinoids to their thermodynamically less stable Z-isomer remains a challenge. In this report, we present a photochemical approach for the catalytic Z-isomerization of retinoids using monochromatic wavelength UV irradiation treatment. We have developed a straightforward approach for the synthesis of Z-retinoids in high yield, overcoming common obstacles normally associated with their synthesis. Calculations based on density functional theory (DFT) have allowed us to correlate the experimentally observed Z-isomer distribution of retinoids with the energies of chemically important intermediates, which include ground- and excited-state potential energy surfaces. We also demonstrate the application of the current method by synthesizing gram-scale quantities of 9-cis-retinyl acetate 9Z-a. Operational simplicity and gram-scale ability make this chemistry a very practical solution to the problem of Z-isomer retinoid synthesis
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