Since 1982, phycobiliproteins have served as fluorescent labels in a wide variety of cell and molecule analyses. The
exceptional spectroscopic properties of these labels include very high absorbance coefficients and quantum yields, and
large Stokes shifts. The spectroscopic diversity of these reagents is restricted to a subset of naturally occurring
phycobiliproteins with stable assembly states in vitro, whose target specificity is generated by chemical conjugation to
proteins or small molecules. The latter step generates heterogeneity. These limitations have been overcome by
expressing various recombinant phycobiliprotein constructs in the cyanobacterium Anabaena sp. PCC7120. Modular
recombinant phycobiliprotein-based labels were constructed with some or all of the following features (a) an affinity
purification tag; (b) a stable oligomerization domain (to maintain stable higher order assemblies of the phycobiliprotein
monomers at very low protein concentration); (c) a biospecific recognition domain. Such phycobiliprotein constructs are
readily purified from crude cell extracts by affinity chromatography and used directly as fluorescent labels. To generate
constructs for intracellular in vivo labeling, the entire pathways for the biosynthesis of the His-tagged holo-
α (phycocyanobilin-bearing) subunit of phycocyanin (emission max. 641 nm) and of the His-tagged holo-α
(phycobiliviolin-bearing) subunit of phycoerythrocyanin (emission max. 582 nm) were reconstituted in Escherichia coli.
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