15 research outputs found
Divergent Behavior of Glycosylated Threonine and Serine Derivatives in Solid Phase Peptide Synthesis
Solid phase peptide coupling of glycosylated threonine derivatives was systematically evaluated. In contrast to glycosylated serine derivatives which are highly prone to epimerization, glycosylated threonine derivatives produce only negligible amounts of epimerization. Under forcing conditions, glycosylated threonine analogs undergo Ī²-elimination, rather than epimerization. Mechanistic studies and molecular modeling were used to understand the origin of the differences in reactivity
Conformational Determinants of the Activity of Antiproliferative Factor Glycopeptide
The antiproliferative factor (APF)
involved in interstitial cystitis
is a glycosylated nonapeptide (TVPAAVVVA) containing a sialylated
core 1 Ī±-O-disaccharide linked to the N-terminal threonine.
The chemical structure of APF was deduced using spectroscopic techniques
and confirmed using total synthesis. The synthetic APF provided a
platform to study amino acid modifications and their effect on APF
activity, based on which a structureāactivity relationship
(SAR) for APF activity was previously proposed. However, this SAR
model could not explain the change in activity associated with minor
alterations in the peptide sequence. Presented is computational analysis
of 14 APF derivatives to identify structural trends from which a more
detailed SAR is obtained. The APF activity is found to be dictated
by the close interplay between carbohydrateāpeptide and peptideāpeptide
interactions. The former involves hydrogen bond and hydrophobic interactions,
and the latter is dominated by hydrophobic interactions. The highly
flexible hydrophobic peptide adopts collapsed conformations separated
by low energy barriers. APF activity correlates with hydrophobic clustering
associated with amino acids 4<b>A</b>, 6<b>V</b>, and
8<b>V</b>. Peptide conformations are highly sensitive to single
point mutations, which explain the experimental trends. The presented
SAR will act as a guide for lead optimization of more potent APF analogues
of potential therapeutic utility
GalNAc-Tyrosine Is a Ligand of Plant Lectins, Antibodies, and Human and Murine Macrophage Galactose-Type Lectins
In
2011, a new type of protein <i>O</i>-glycosylation
was discovered in which <i>N</i>-acetylgalactosamine is
attached to the side chain of tyrosine (GalNAc-Tyr). While present
on dozens of proteins, the biological roles of GalNAc-Tyr are unknown.
To gain insight into this new type of modification, we synthesized
a group of GalNAc-Tyr glycopeptides, constructed microarrays, and
evaluated potential recognition of GalNAc-Tyr by a series of glycan-binding
proteins. Through a series of >150 microarray experiments, we assessed
binding properties of a variety of plant lectins, monoclonal antibodies,
and endogenous lectins. VVL, HPA, and SBA were all found to bind tightly
to GalNAc-Tyr, and several Tn binding antibodies and blood group A
antibodies were found to cross-react with GalNAc-Tyr. Thus, detection
of GalNAc-Tyr modified proteins is an important consideration when
analyzing results from these reagents. Additionally, we evaluated
potential recognition by two mammalian lectins, human (hMGL) and murine
(mMGL-2) macrophage galactose type C-type lectins. Both hMGL and mMGL-2
bound tightly to GalNAc-Tyr determinants. The apparent <i>K</i><sub>d</sub> values (ā¼1ā40 nM) were on par with some
of the best known ligands for MGL, such as the Tn antigen. hMGL also
bound the natural beta-amyloid peptide containing a GalNAc-Tyr epitope.
STD NMR experiments provided structural insights into the molecular
basis of recognition. Finally, GalNAc-Tyr was selectively captured
by mMGL-2 positive dendritic cells. These results provide the first
evidence that GalNAc-Tyr modified proteins and/or peptides may be
ligands for hMGL and mMGL-2 and offer unique structures for the design
of MGL targeting agents
Side-by-side view of () T7T8 (with residues labeled) and () the native dodecamer to illustrate the position and potential contacts of the additional atoms in the bicyclo3
1D NMR spectra of () T7 and () T7T8 imino regions at 200 Ī¼M strand concentration
<p><b>Copyright information:</b></p><p>Taken from "Biophysical studies of DNA modified with conformationally constrained nucleotides: comparison of 2ā²- (north) and 3ā²- (south) ālockedā templates"</p><p>Nucleic Acids Research 2007;35(6):1978-1991.</p><p>Published online 6 Mar 2007</p><p>PMCID:PMC1895885.</p><p>Ā© 2007 The Author(s)</p> Additional peaks that were not observed in 1ā2āmM solutions are indicated with arrows
Changes in the chemical shift of the imino protons between the -MCT- and -MCT- modified ODN species and the native DD at 5Ā°C () and at 25Ā°C ()
<p><b>Copyright information:</b></p><p>Taken from "Biophysical studies of DNA modified with conformationally constrained nucleotides: comparison of 2ā²- (north) and 3ā²- (south) ālockedā templates"</p><p>Nucleic Acids Research 2007;35(6):1978-1991.</p><p>Published online 6 Mar 2007</p><p>PMCID:PMC1895885.</p><p>Ā© 2007 The Author(s)</p
NMR spectra of the imino H region of the -MCT modified oligos at various temperatures
<p><b>Copyright information:</b></p><p>Taken from "Biophysical studies of DNA modified with conformationally constrained nucleotides: comparison of 2ā²- (north) and 3ā²- (south) ālockedā templates"</p><p>Nucleic Acids Research 2007;35(6):1978-1991.</p><p>Published online 6 Mar 2007</p><p>PMCID:PMC1895885.</p><p>Ā© 2007 The Author(s)</p> The assignments are as follows: filled diamond T8, filled square T7, filled triangle G2, filled circle G4, star G10 and double dagger, G12
Changes in the chemical shift of select non-exchangeable protons between the -MCT- () and -MCT- () modified ODN species and the native DD at 25Ā°C
<p><b>Copyright information:</b></p><p>Taken from "Biophysical studies of DNA modified with conformationally constrained nucleotides: comparison of 2ā²- (north) and 3ā²- (south) ālockedā templates"</p><p>Nucleic Acids Research 2007;35(6):1978-1991.</p><p>Published online 6 Mar 2007</p><p>PMCID:PMC1895885.</p><p>Ā© 2007 The Author(s)</p
Experimental and Chemoinformatics Study of Tautomerism in a Database of Commercially Available Screening Samples
We
investigated how many cases of the same chemical sold as different
products (at possibly different prices) occurred in a prototypical
large aggregated database and simultaneously tested the tautomerism
definitions in the chemoinformatics toolkit CACTVS. We applied the
standard CACTVS tautomeric transforms plus a set of recently developed
ringāchain transforms to the Aldrich Market Select (AMS) database
of 6 million screening samples and building blocks. In 30āÆ000
cases, two or more AMS products were found to be just different tautomeric
forms of the same compound. We purchased and analyzed 166 such tautomer
pairs and triplets by <sup>1</sup>H and <sup>13</sup>C NMR to determine
whether the CACTVS transforms accurately predicted what is the same
āstuff in the bottleā. Essentially all prototropic transforms
with examples in the AMS were confirmed. Some of the ringāchain
transforms were found to be too āaggressiveā, i.e. to
equate structures with one another that were different compounds
Glycoamino Acid Analogues of the ThomsenāFriedenreich Tumor-Associated Carbohydrate Antigen: Synthesis and Evaluation of Novel Antiproliferative Factor Glycopeptides
Glycoamino
acid analogues of the ThomsenāFriedenreich antigen
disaccharide, where the 4ā² and 4ā³ hydroxyl groups were
substituted with fluorine or hydrogen, were synthesized and incorporated
into the asialylated antiproliferative factor (<i>as</i>-APF), a biologically active form of APF, a glycopeptide found in
the urine of patients with interstitial cystitis. Various strategies
were employed to incorporate the fluorine atom at the 4-positions
of either the galactose or <i>N</i>-acetylgalactosamine
unit of the disaccharide antigen, based on stereochemistry and reactivity.
These glycopeptides were evaluated in antiproliferative assays on
both primary normal bladder epithelial cells and T24 bladder carcinoma
cells. Unlike many previously published substitutions to APF, mono-4ā²-fluorination
of the GalNAc residue did not affect the activity, whereas fluoro-derivatives
of the galactose 4ā³-position or both 4ā² and 4ā³
hydroxyls showed a reduced potency relative to the monosubstituted
GalNAc derivative. A fourth compound where the 4ā³ position
of galactose was deoxygenated showed a lower potency than the parent
and monosubstituted compounds. These results suggest that specific
substitutions in the sugar moieties in the APF can be tolerated, and
the glycomimetic design of APF analogues can include fluorine in the
GalNAc sugar of the disaccharide