The structure of a protein complex
needs to be controlled appropriately
to maximize its functions. Herein, we report the linear polymerization
of bacterial alkaline phosphatase (BAP) through the site-specific
cross-linking reaction catalyzed by Trametes sp.
laccase (TL). We introduced a peptide loop containing a tyrosine (Y-Loop)
to BAP, and the Y-Looped BAP was treated with TL. The Y-Looped BAP
formed linear polymers, whereas BAP fused with a C-terminal peptide containing a tyrosine (Y-tag) showed an irregular
shape after TL treatment. The sterically confined structure of the
Y-Loop could be responsible
for the formation of linear BAP polymers. TL-catalyzed copolymerization
of Y-Looped BAP and a Y-tagged chimeric antibody-binding protein,
pG2pA-Y, resulted in the formation of linear bifunctional
protein copolymers that could be employed as protein probes in an
enzyme-linked immunosorbent assay (ELISA). Copolymers comprising Y-Looped
BAP and pG2pA-Y at a molar ratio of 100:1 exhibited the
highest signal in the ELISA with 26- and 20-fold higher than a genetically
fused chimeric protein, BAP-pG2pA-Y, and its polymeric
form, respectively. This result revealed that the morphology of the
copolymers was the most critical feature to improve the functionality
of the protein polymers as detection probes, not only for immunoassays
but also for other diagnostic applications.