Purification of the yellow fluorescent protein from vibrio fischeri and identity of the flavin chromophore

Abstract

A low molecular weight protein (∼ 25,000 D) exhibiting a yellow fluorescence emission peaking at ∼ 540 nm was isolated from Vibrio fischeri (strain Y-1) and purified to apparent homogeneity. FMN is the chromophore, but it exhibits marked red shifts in both the absorption (λ max = 380, 460 nm) and the fluorescence emission. When added to purified luciferase from the same strain, which itself catalyzes an emission of blue-green light (λ max ∼ 495 nm), this protein induces a bright yellow luminescence (λ max ∼ 540 nm); this corresponds to the emission of the Y-1 strain in vivo . This yellow bioluminescence emission is thus ascribed to the interaction of these two proteins, and to the excitation of the singlet FMN bound to this fluorescent protein.