Abstract
Dark-operative protochlorophyllide (Pchlide) reductase (DPOR) is a nitrogenase-like enzyme catalyzing the D-ring reduction of Pchlide in the penultimate step of chlorophyll biosynthesis in most oxygenic photosynthetic organisms. DPOR is the determinant enzyme for greening in the dark of the photosynthetic organisms. How the oxygen-sensitive enzyme operates in oxygenic photosynthetic cells remains unsolved. Here we report the functional expression of the cyanobacterial DPOR catalytic component, NB-protein (ChlN-ChlB), in E. coli. The chlN and chlB genes from the cyanobacterium Leptolyngbya boryana (formerly Plectonema boryanum) were artificially connected to form a small operon, chlN-chlB, in the overexpression plasmid. ChlN protein with Sterp-tag was co-purified with ChlB by an affinity column from the soluble fraction of E. coli. The addition of thus purified NB-protein to a crude extract of the LPOR-lacking mutant of L. boryana obviously stimulated the DPOR activity, indicating that an active form of NB-protein was expressed in E. coli.
