Single amino acid changes affect CD33 sialoglycan binding. (A) Sialoglycan-binding profile of purified, soluble, recombinant CD33 proteins was determined using a sialoglycan microarray containing defined, chemically synthesized glycans (experiment schematic shown in B). Nonsialylated, Neu5Ac- (indicated in purple on the left) and Neu5Gc- (indicated in blue) terminating glycans were grouped together in the heatmap as shown in the left. Each column indicates the binding profile of the protein indicated on the top and each row represent a distinct glycan. Blue indicates no binding and red indicates very strong binding preferences characterized by an average relative fluorescence unit (RFU) of more than 90th percentile. The result of the heatmap is summarized in the cartoon (C) with the colored diamonds corresponding to the termination Sia and supplementary figure S2, Supplementary Material online. The names of the individual glycans are presented in supplementary file S1, Supplementary Material online.