Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Pivot Concepts:
Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Target Concepts:
Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Query: EC:1.10.3.2 (
laccase
)
4,656
document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)
The site-specific cross-linking of functional proteins creates macromolecular assemblies that exhibit unique biochemical and/or physicochemical properties. Herein, we explored the potential of
laccase
as a biocatalyst for the site-specific cross-linking of tyrosine-tagged proteins. Trametes sp.
laccase
(TL) was selected as the cross-linking catalyst, and Escherichia coli alkaline phosphatase (
BAP
) and antibody-binding proteins (pG
2
pAs) were employed as model proteins. The protein models were genetically fused to a peptide tag containing a tyrosine residue (Y-tag) at the N- and/or C-termini. Proteins without Y-tags were used as controls. The Y-tagged proteins could be recognized by TL as macromolecular substrates, leading to the oxidative formation of protein polymers, whereas no polymerization was observed with intact
BAP
or pG
2
pA. The TL-catalyzed cross-linking of Y-tagged proteins proceeded at a relatively high pH in comparison with that of small phenolic substrates. Co-polymers of
BAP
and pG
2
pA were able to be prepared by mixing the aqueous solution of each component in the presence of TL. A combination of bis-Y-tagged pG
2
pA (Y-pG
2
pA-Y) and Y-tagged
BAP
(
BAP
-Y) yielded functional co-polymers compatible with enzyme-linked immunosorbent assay (ELISA). The detection limit of the ELISA of ovalbumin with anti-OVA IgG depended on the molar ratio of
BAP
-Y and Y-pG
2
pA-Y in the TL-catalyzed cross-linking reaction. A high molar ratio of
BAP
-Y to Y-pG
2
pA-Y (75:1) resulted in the highest absorbance in the ELISA. The results suggested that the formation of a bifunctional protein polymer with a high molar ratio of signaling unit to antibody-binding unit gave better performance in antigen detection than using lower ratios.
...
PMID:Laccase-catalyzed bioconjugation of tyrosine-tagged functional proteins. 2990 78
