Streptococcus gordonii has an extracellular glucosyltransferase (GTF) that polymerizes the glucose moiety of sucrose to form both water-soluble and water-insoluble glucans. Whereas multiple gtf genes have been identified in strains of mutans streptococci and Streptococcus salivarius, a single gene, designated gtfG, encodes the GTF of S. gordonii Challis. gtfG is also unique among the characterized gtfs in that it has a described regulatory determinant, rgg. Furthermore, the GTF activity in S. gordonii undergoes reversible phase variation between high and low levels. In order to gain insight into this novel GTF system, the nucleotide sequence of gtfG was determined and found to consist of a 4,734 base pair open reading frame encoding a protein with a deduced molecular weight of ca. 174,000. gtfG was similar to other sequenced gtfs with a conserved signal sequence followed by a ca. 600-bp region distinctive for gtfG, a conserved region encoding a putative catalytic active site and a series of six direct repeats in the carboxyl terminal region implicated in glucan binding. Although comparison of gtfG to other gtfs did not show a basis for the primer-independence of the encoded enzyme or the nature of the glucan products, the gtfG sequence data provide an important basis for further studies of these enzymes.