Enzymatic glucosylation with glycosyltransferases can be used to regulate the water solubility of aglycone. The drawback of this process is the demand of UDP-glucose as a sugar donor. We made an in-frame fusion of the flavonoid O-glucosyltransferase (OsUGT-3) and sucrose synthase (AtSUS) genes. The resulting fusion protein, OsUGT3-AtSUS, was expressed in E. coli and purified. When sucrose and UDP were supplied, the fusion protein was able to convert quercetin into quercetin O-glucoside without the addition of UDP-glucose. In addition, UDP-glucose was recycled when sucrose was added to the reaction mixture. This fusion protein is useful for the enzymatic production of flavonoid O-glucosides.