A lectin with specificity toward mannose, glucose, and rhamnose has been isolated from the legumes of the knife bean Canavalia gladiata. The lectin is composed of two identical 30-kDa subunits with substantial N-terminal sequence similarity to Concanavalin A (Con A). It was purified by affinity chromatography on Affi-gel blue gel, ion exchange chromatography on Mono S, and gel filtration by fast protein liquid chromatography on Superdex 200. Compared with Con A, knife bean lectin started to stimulate [methyl-(3)H]thymidine uptake by mouse splenocytes at a lower concentration, and more potently inhibited proliferation of L1210 leukemia cells. In contrast to Con A, the mitogenic activity of knife bean lectin toward mouse splenocytes, but not its antiproliferative activity toward tumor cells, could be abrogated by 250 mM glucose. Both mitogenic and antiproliferative activities of Con A were abolished by glucose. The lectin inhibited HIV-1 reverse transcriptase with an IC(50) of 35 microM and cell-free translation in a rabbit reticulocyte lysate system with an IC(50) of 2.08 microM. The lectin did not exhibit antifungal activity.