Pseudomonas aeruginosaA‐band lipopolysaccharide (LPS) molecule has an O‐polysaccharide region composed of trisaccharide repeat units of α1 → 2, α1 → 3, α1 → 3 linked D‐rhamnose (Rha). The A‐band polysaccharide is assembled by the α‐ D‐rhamnosyltransferases, WbpX, WbpY and WbpZ. WbpZ probably transfers the first Rha residue onto the A‐band accepting molecule, while WbpY and WbpX subsequently transfer two α1 → 3 linked Rha residues and one α1 → 2 linked Rha respectively. The last two transferases are predicted to be processive, alternating in their activities to complete the A‐band polymer. The genes coding for these transferases were identified at the 3′ end of the A‐band biosynthetic cluster. Two additional genes, psecoAand uvrD, border the 3′ end of the cluster and are predicted to encode a co‐enzyme A transferase and a DNA helicase II enzyme respectively. Chromosomal wbpX, wbpYand wbpZmutants were generated, and Western immunoblot analysis demonstrates that these mutants are unable to synthesize A‐band LPS, while B‐band synthesis is unaffected. WbpL, a transferase encoded within the B‐band biosynthetic cluster, was previously proposed to initiate B‐band biosynthesis through the addition of Fuc2NAc (2‐acetamido‐2,6‐dideoxy‐ D‐galactose) to undecaprenol phosphate (Und‐P). In this study, chromosomal wbpLmutants were generated that did not express A band or B band, indicating that WbpL initiates the synthesis of both LPS molecules. Cross‐complementation experiments using WbpL and its homologue, Escherichia coliWecA, demonstrates that WbpL is bifunctional, initiating B‐band synthesis with a Fuc2NAc residue and A‐band synthesis with either a GlcNAc ( N‐acetylglucosamine) or GalNAc ( N‐acetylgalactosamine) residue. These data indicate that A‐band polysaccharide assembly requires four glycosyltransferases, one of which is necessary for initiating both A‐band and B‐band LPS synthesis.