3 research outputs found
Chemoenzymatic Probes for Detecting and Imaging Fucose-Ī±(1-2)-galactose Glycan Biomarkers
The disaccharide motif fucose-Ī±Ā(1-2)-galactose
(FucĪ±Ā(1-2)ĀGal)
is involved in many important physiological processes, such as learning
and memory, inflammation, asthma, and tumorigenesis. However, the
size and structural complexity of FucĪ±Ā(1-2)ĀGal-containing glycans
have posed a significant challenge to their detection. We report a
new chemoenzymatic strategy for the rapid, sensitive detection of
FucĪ±Ā(1-2)ĀGal glycans. We demonstrate that the approach is highly
selective for the FucĪ±Ā(1-2)ĀGal motif, detects a variety of complex
glycans and glycoproteins, and can be used to profile the relative
abundance of the motif on live cells, discriminating malignant from
normal cells. This approach represents a new potential strategy for
biomarker detection and expands the technologies available for understanding
the roles of this important class of carbohydrates in physiology and
disease
Identification of the Plasticity-Relevant Fucose-Ī±(1ā2)-Galactose Proteome from the Mouse Olfactory Bulb
Fucose-Ī±(1ā2)-galactose [FucĪ±(1ā2)Gal] sugars have been implicated in the molecular mechanisms that underlie neuronal development, learning, and memory. However, an understanding of their precise roles has been hampered by a lack of information regarding FucĪ±(1ā2)Gal glycoproteins. Here, we report the first proteomic studies of this plasticity-relevant epitope. We identify five classes of putative FucĪ±(1ā2)Gal glycoproteins: cell adhesion molecules, ion channels and solute carriers/transporters, ATP-binding proteins, synaptic vesicle-associated proteins, and mitochondrial proteins. In addition, we show that FucĪ±(1ā2)Gal glycoproteins are enriched in the developing mouse olfactory bulb (OB) and exhibit a distinct spatiotemporal expression that is consistent with the presence of a āglycocodeā to help direct olfactory sensory neuron (OSN) axonal pathfinding. We find that expression of FucĪ±(1ā2)Gal sugars in the OB is regulated by the Ī±(1ā2)fucosyltransferase FUT1. FUT1-deficient mice exhibit developmental defects, including fewer and smaller glomeruli and a thinner olfactory nerve layer, suggesting that fucosylation contributes to OB development. Our findings significantly expand the number of FucĪ±(1ā2)Gal glycoproteins and provide new insights into the molecular mechanisms by which fucosyl sugars contribute to neuronal processes
Cell-Specific Chemical Delivery Using a Selective NitroreductaseāNitroaryl Pair
<p>The utility of<b> </b>small molecules to probe or perturb biological systems is limited by the lack of cell-specificity. āMaskingā the activity of small molecules using a general chemical modification and āunmaskingā it only within target cells could overcome this limitation. To this end, we have developed a selective enzymeāsubstrate pair consisting of engineered variants of <i>E. coli</i> nitroreductase (NTR) and a 2ānitro-<i>N</i>-methylimidazolyl (NM) masking group. To discover and optimize this NTRāNM system, we synthesized a series of fluorogenic substrates containing different nitroaromatic masking groups, confirmed their stability in cells, and identified the best substrate for NTR. We then engineered the enzyme for improved activity in mammalian cells, ultimately yielding an enzyme variant (enhanced NTR, or eNTR) that possesses up to 100-fold increased activity over wild-type NTR. These improved NTR enzymes combined with the optimal NM masking group enable rapid, selective unmasking of dyes, indicators, and drugs to genetically defined populations of cells.</p