Gonococcal lipooligosaccharide (LOS) is a major virulence factor. Its abundance and surface-exposure make LOS an excellent target for vaccines and immunotherapeutics. A well-studied monoclonal antibody (mAb) known as 2C7 recognizes a majority of clinical isolates and mediates complement-dependent bactericidal activity. mAb 2C7 was humanized by CDR grafting and its Fc was engineered to enhance complement activation. Humanized 2C7 decreased gonococcal burden in a mouse vaginal colonization model. To identify new human anti-LOS mAbs, we immunized transgenic mice that expressed the variable regions of human IgG heavy and light chains with LOS and heat-killed gonococci. One of the mAbs, designated 3-19-23, bound to an LOS structure distinct from the 2C7 epitope. Both mAbs required expression of HepII lactose (beta-chain) to bind to LOS. mAb 2C7, but not 3-19-23 bound to LOS structures with glycan extensions beyond HepI (alpha-chain) lactose. While mAb 2C7 killed five and mAb 3-19-23 killed eight of ten WHO reference strains tested, nine strains were killed in a complement-dependent manner by at least one of the two mAbs, demonstrating complementary functional activities. To define the structural basis for the specificities of the two mAbs, we determined high-resolution crystal structures of Fabs 2C7 and 3-19-23, each bound to oligosaccharide (OS) extracted and purified from strain 15253 (expresses lactose from alpha- and beta-chains) at 1.6 and 1.3-Å resolution, respectively. The two Fabs bound OS in distinct orientations: Fab 2C7 recognized the beta and gamma glycan chains, whereas Fab 3-19-23 recognized the alpha and beta chains. The alpha chain was more exposed to solvent in the Fab 2C7-OS complex than in the 3-19-23 complex, which explains the greater tolerance of mAb 2C7 for binding LOS with alpha chain extensions beyond lactose. The beta chain was more tightly packed and less exposed to solvent in the Fab 2C7 complex than in the 3-19-23 complex. These features provide a structural basis for why sialylation of the beta chain of LOS decreases the bactericidal activity of mAb 2C7, but not 3-19-23. Collectively, our data provide a strong rationale for the development of vaccines and mAbs targeting LOS to combat antimicrobial-resistant gonorrhea.