In pea membranes, UDP[¹⁴C]Glc glycosylates a ~40-kDa polypeptide doublet. This label rapidly disappears if excess unlabeled UDP-Glc, or UDP, is added, indicating that the glycosylation is reversible, and suggesting that the glycosylated polypeptides might be intermediates in a glycosyl transfer reaction. Glycosylation of the doublet requires a divalent cation, the effective ions being the same (except for Zn²⁺) as those that activate Golgi-localized β-glucan synthase (GS-I) activity. Treatments that inhibit GS-I also inhibit doublet glycosylation. The doublet is associated with Golgi (and to a minor extent with plasma) membranes and occurs also in the soluble fraction. The Golgi-bound doublet may be a component of the GS-I system. Immunological, inactivation, and fractionation evidence indicates that at least one other polypeptide is required in GS-I activity.