Characterization of R-phycocyanin. Chromophore content of R-phycocyanin and C-phycoerythrin.

摘要: R-phycocyanin was purified from two independent isolates of the unicellular red alga Porphyridium cruentum. At pH 7.0 protein sediments as a single component with s 20,w 5.98 S (at 2 mg/ml, gamma/2=0.02). Over concentration range 0.2 to 0.5 mg/ml (gamma/2=0.16), sedimentation equilibrium gave molecular weight 103,000 +/- 6,000 no evidence heterogeneity. In common C-phydocyanins, consists alpha and beta subunits weights 18,200 20,500, determined by electrophoresis in sodium dodecyl sulfate-polyacrylamide gels. Isoelectric focusing polyacrylamide gels resolves bands, blue pI 5.2), purple 5.3), believed correspond subunits, respectively. The native precipitin band when tested against homologous antiserum Ouchterlony double diffusion technique. No cross-reaction observed allophycocyanin same organism. absorption spectrum trimeric R-physocyanin at exhibited epsilomN (555 nm) 1.51 x 10(5) M(-1) cm(-1), epsilonM (618 2.55 A 1% 1cm 70.0. circular dichroism characterized following ellipticity maxima deg cm2 per dmol 10(-5): [theta]311 = -2.36, [theta]343 -3.27, [theta]552 4.67, [theta]627 6.27. All these values were based on an alphabeta 36,3000, calculated amino acid composition. To permit quantitative estimation chromophore composition R-phycocyanin, properties Aphanocapsa sp. C-phycoerythrin determined. 7.0, (562 4.88 127, 38,400 molar extinction coefficients for polypeptide-bound phycoerythrobilin denatured 8 M urea 1.9, assumption that each unit contains six such chromophores. analogous data phycocyanobilin available earlier study (Glazer, A.N., Fang, S. (1973) J. Biol. Chem. 248, 659-662). curve fitted high precision theoretical mixture one chromophore. analyses its separated demonstrated 1:1 stoichiometry protein. spectra isolated consistent conclusion subunit carries chromophore, while are bound subunit...