Chromophore topography and secondary structure of 124-kilodalton Avena phytochrome probed by zinc-induced chromophore modification

Sommer, Debbie; Song, Pill Soon

doi:10.1021/bi00459a040

Cited by 37 publications

(36 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Immediately after the end of the irradiation, 5 M HC1 was added to lower the pH to 9.0. The alkaline treatment at pH 11.0 yielded a stable bleached spectrum at pH 9.0 similar to those of soyasaponin-treated (Konomi et al, 1982), tryptic digestion-treated (Yamamoto & Furuya, 1983), and zinciontreated (Sommer & Song, 1990) phytochromes, although alkaline treatment lower than pH 11 .O showed reversion to PI at pH 9.0.…”

mentioning

confidence: 74%

“…A model of Pbl was prepared by alkaline treatment, since the addition of bleaching reagents such as zinc ions (Sommer & Song, 1990) or soyasaponin (Konomi et al, 1982) to the highly concentrated phytochrome solutions induced some precipitation. In brief, large phytochrome precipitated with ammonium sulfate was resuspended in 50 mM CHES and 1 mM Na2EDTA at pH 9.0, and then its pH was raised to 11.0 by adding 5 M NaOH.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Resonance Raman spectra of the intermediates in phototransformation of large phytochrome: Deprotonation of the chromophore in the bleached intermediate

Mizutani¹,

Tokutomi²,

Kitagawa³

1994

Biochemistry

View full text Add to dashboard Cite

Resonance Raman (RR) scattering from type A large phytochrome of pea was measured at cryogenic as well as ambient temperatures to determine an intermediate in which deprotonation of the chromophore takes place. The RR bands of the red-absorbing (Pr) and far-red-absorbing forms (Pfr) of large pea phytochromes at ambient temperatures are almost the same in their frequencies as those of the intact form reported previously (Mizutani et al., 1991). The RR spectrum of large phytochrome excited at 364 nm at -120 degrees C, where Pr and a photointermediate, I700 (= lumi-R), are trapped, showed a strong band at 1625 cm-1 with a shoulder at 1648 cm-1 in the C C=C stretching region. The shoulder disappeared, and a new band appeared at 1597 cm-1 upon raising the temperature to -80 degrees C, where transformation from I700 to meta-Ra proceeds. The RR spectra remained unchanged until -10 degrees C, indicating that the RR spectra of meta-Rb and meta-Rc are close to that of meta-Ra, and we call them comprehensively the bleached intermediate, Ibl. At ambient temperatures where photo-steady-states among a few species are attained, strong RR bands were observed at 1625 and 1599 cm-1 upon excitation at 364 nm under simultaneous far-red illumination, and the 1599-cm-1 band was appreciably intensified under simultaneous red- instead of far-red illumination. By comparison of these spectra with those at low temperatures, the 1625- and 1599-cm-1 bands were reasonably assigned to Pr and Ibl, respectively. A chemically prepared model of the bleached form, Pbl, also gave a prominent band at 1599 cm-1.(ABSTRACT TRUNCATED AT 250 WORDS)

show abstract

mentioning

confidence: 74%

mentioning

confidence: 99%

Resonance Raman spectra of the intermediates in phototransformation of large phytochrome: Deprotonation of the chromophore in the bleached intermediate

Mizutani¹,

Tokutomi²,

Kitagawa³

1994

Biochemistry

View full text Add to dashboard Cite

show abstract

“…The ratio of [θ] 190 /[θ] 220 values indicated that the predominant secondary structural component in all Cph1 proteins analyzed was the R-helical conformation (Table 2), as in plant phytochromes (33,34). The R-helical content of the apoprotein was calculated to be 37%.…”

Section: Expression and In Vitro Reconstitution Of The Cph1mentioning

confidence: 99%

Chromophore−Apoprotein Interactions in Synechocystis sp. PCC6803 Phytochrome Cph1

et al. 2000

View full text Add to dashboard Cite

The secondary, tertiary, and quaternary structures of the Synechocystis Cph1 phytochrome were investigated by absorption and circular dichroism spectroscopy, size exclusion chromatography, and limited proteolysis. The Cph1 protein was coexpressed with a bacterial thioredoxin in Escherichia coli, reconstituted in vitro with tetrapyrrole chromophores, and purified by chitin affinity chromatography. The resultant Cph1 holoproteins were essentially pure and had the specific absorbance ratio (SAR) of 0.8-0.9. Circular dichroism spectroscopy and limited proteolysis showed that the chromophore binding induced marked conformational changes in the Cph1 protein. The alpha-helical content increased to 42-44% in the holoproteins from 37% in the apoprotein. However, no significant difference in the secondary structure was detected between the Pr and Pfr forms. The tertiary structure of the Cph1 apoprotein appeared to be relatively flexible but became more compact and resistant to tryptic digestion upon chromophore binding. Interestingly, a small chromopeptide of about 30 kDa was still predominant even after longer tryptic digestion. The N-terminal location of this chromopeptide was confirmed by expression in E. coli and in vitro reconstitution with chromophores of the 32.5 kDa N-terminal fragment of the Cph1 protein. This chromopeptide was fully photoreversible with the spectral characteristic similar to that of the full-size Cph1 protein. The Cph1 protein forms dimers through the C-terminal region. These results suggest that the prokaryotic Cph1 phytochrome shares the structural and conformational characteristics of plant phytochromes, such as the two-domain structure consisting of the relatively compact N-terminal and the relatively flexible C-terminal regions, in addition to the chromophore-induced conformational changes.

show abstract

“…A specific conformational change occurs along the amino terminal chain of phyA upon its phototransfonnation from the Pr to the Pfr form (Chai et al 1987;Vierstra et al 1987;Sommer & Song 1990). To test the reliability of the circular dichroism (CD) spectroscopy, three different CD analysis methods have been used to determine the secondary structure of pea phyA in both the Pr and the Pfr forms (Deforce et al 1994).…”

Section: Light-induced Conformational Changesmentioning

confidence: 99%

“…To test the reliability of the circular dichroism (CD) spectroscopy, three different CD analysis methods have been used to determine the secondary structure of pea phyA in both the Pr and the Pfr forms (Deforce et al 1994). We showed that the secondary structure of dicot pea phyA is very similar to that of monocot oat phyA (Sommer & Song 1990), interdependent of the method of CD analysis (Deforce et al 1994). As with oat phyA, an increase in a-helix of the apoprotein polypeptide takes place when the Pr form is phototransfonned to the Pfr form.…”

Section: Light-induced Conformational Changesmentioning

confidence: 99%

Chromophore: apoprotein interactions in phytochrome A*

Song

Park

Furuya

1997

Plant Cell & Environment

View full text Add to dashboard Cite

Pbytocbromes are molecular ligbt switcbes by virtue of their pbotocbromic red/far-red reversibility. The His-324 residue next to the chromophore-Iinked Cys-323 plays a critical role in conferring photochromism to the tetrapyrrole chromophore in native phytochrome A. The chromophore appears to be enclosed between the amphiphilic a-helical chains in a hydrophobie pocket. The absorbance maxima of both the Pr and the Pfr forms of pea phytochrome A are blue-shifted by 10 and 20 nm, respectively, upon C-terminal truncation. We speculate that the quaternary structure of the phytochrome A molecule involves some interactions of the C-terminal half with the chromophore domain. The Pfr conformation of phytochrome includes an amphiphilic ahelix of the amino terminal chain, which occurs in 113 ms after picosecond photoisomerization of the Pr form. Compared to a-helical folding, unfolding of the a-helix occurs faster in about 310 fis upon phototransformation of the Pfr form of phytochrome A. The photochromic transformation of phytochrome A modulates protein kinase-catalysed phosphorylation sites in vivo and in vitro, but only a subtle local change in conformation is detectable in the phosphorylated phytochromes. This suggests that the post-transiational modification serves as a surface label, rather than a transducer-activating trigger, for the recognition of a putative phytochrome receptor.

show abstract

Chromophore topography and secondary structure of 124-kilodalton Avena phytochrome probed by zinc-induced chromophore modification

Cited by 37 publications

References 34 publications

Resonance Raman spectra of the intermediates in phototransformation of large phytochrome: Deprotonation of the chromophore in the bleached intermediate

Resonance Raman spectra of the intermediates in phototransformation of large phytochrome: Deprotonation of the chromophore in the bleached intermediate

Chromophore−Apoprotein Interactions in Synechocystis sp. PCC6803 Phytochrome Cph1

Chromophore: apoprotein interactions in phytochrome A*

Contact Info

Product

Resources

About