Carbohydrate arrays (glycoarrays) have recently emerged as a high-throughput tool for studying carbohydrate-binding proteins and carbohydrate-processing enzymes. A number of sophisticated array platforms that allow for qualitative and quantitative analysis of carbohydrate binding and modification on the array surface have been developed, including analysis by fluorescence spectroscopy, mass spectrometry and surface plasmon resonance spectroscopy. These platforms, together with examples of biologically-relevant applications are reviewed in this Feature Article.
Plant lectin recognition of glycans was evaluated by SPR imaging using a model array of N-biotinylated aminoethyl glycosides of beta-D-glucose (negative control), alpha-D: -mannose (conA-responsive), beta-D-galactose (RCA(120)-responsive) and N-acetyl-beta-D-: glucosamine (WGA-responsive) printed onto neutravidin-coated gold chips. Selective recognition of the cognate ligand was observed when RCA(120) was passed over the array surface. Limited or no binding was observed for the non-cognate ligands. SPR imaging of an array of 40 sialylated and unsialylated glycans established the binding preference of hSiglec7 for alpha2-8-linked disialic acid structures over alpha2-6-sialyl-LacNAcs, which in turn were recognized and bound with greater affinity than alpha2-3-sialyl-LacNAcs. Affinity binding data could be obtained with as little as 10-20 microg of lectin per experiment. The SPR imaging technique was also able to establish selective binding to the preferred glycan ligand when analyzing crude culture supernatant containing 10-20 microg of recombinant hSiglec7-Fc. Our results show that SPR imaging provides results that are in agreement with those obtained from fluorescence based carbohydrate arrays but with the added advantage of label-free analysis.
Glycoarrays on gold: A designer gold surface incorporating a self-assembled monolayer with weak protein absorption properties has been optimised for rapid display and interrogation of both native and derivatised glycans in array formats. This rapid, facile approach has diverse applications in glycomics, through exploitation of fluorescence, SPR and MALDI-ToF MS detection methods
Play it again, SAM. Peptide substrates immobilised on self‐assembled monolayer (SAM)‐coated gold surfaces were efficiently glycosylated with UDP‐N‐acetyl‐α‐D‐galactosamine:polypeptide N‐acetylgalactosaminyltransferase 2 (ppGalNAcT2) to form the α‐D‐GalNAc threonine/serine linkage. Label‐free analysis of the reactions were performed by using MALDI‐ToF mass spectrometry.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.