Bilin chromophores as reporters of unique protein conformations of phycocyanin 645.
At 45 degrees C, phycocyanin 645 maximally undergoes a reversible and stable conformational change. The change is observed in the visible (chromophore) region of the absorption and circular dichroism (CD) spectra. In the absorption spectrum, the absorbance is lower at 45 degrees C but remains much closer to the normal spectrum than to a strongly denatured spectrum. In the CD, a similar situation exists except that a negative band on the blue edge of the spectrum is much more strongly affected at 45 degrees C than the other bands. On returning to 20 degrees C, all these changes are restored to the original states. The protein is an alpha 2 beta 2 dimer at both 20 and 45 degrees C, and CD in the far-UV shows the identical protein secondary structures at both 20 and 45 degrees C. Fluorescence studies show that energy transfer occurs at both temperatures. At 50 degrees C the results are saliently different as the secondary structure changes and the spectral changes are mostly irreversible. At 50 degrees C, some monomers (alpha beta) are produced, and these monomers are very unstable at that temperature, resulting in the formation of some fully denatured polypeptides. Stable monomers can be produced at 20 degrees C and have visible absorption and CD spectra identical to the dimer at 45 degrees C. Therefore, the chromophores are reporting a tertiary conformational change at 45 degrees C, in which the two halves of the dimer each assume a monomer-like conformation prior to dissociating. These results are compared with a hypothesis for the chromophore topography, and the CD change at the blue edge of the spectra may result from the separation at 45 degrees C of a chromophore pair.
MacColl
R
R
Wadsworth Center, New York State Department of Health, Albany 12201-0509, USA. robert.maccoll@wadsworth.org
Kapoor
S
S
Montellese
D R
DR
Kukadia
S
S
Eisele
L E
LE
D001654:Bile Pigments / Q000737:chemistry*; D002942:Circular Dichroism; D006358:Hot Temperature; D010798:Phycocyanin / Q000737:chemistry*; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet
D001654:Bile Pigments; C039637:phycocyanin 645; D010798:Phycocyanin