Small-angle X-ray scattering studies on the macromolecular structure of the red-light-absorbing form of 121 kDa pea phytochrome and its 114 kDa chromopeptide

Satoru Tokutomi, Mikio Kataoka, Jun Sakai, Masayoshi Nakasako, Fumio Tokunaga, Mitsuo Tasumi, Masaki Furuya

Research output: Contribution to journalArticle

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Abstract

The macromolecular structure of 121 kDa pea phytochrome and its 114 kDa chromopeptide was studied by small-angle X-ray scattering (SAXS) and steric exclusion column chromatography. The molecular mass of the 114 kDa chromopeptide in the red-light-absorbing form (Pr) was determined to be 228 kDa in 100 mM potassium phosphate and 1 mM EDTA (pH 7.8) by SAXS, indicating that the chromopeptide is a dimer. The radius of gyration (Rg) of the chromopeptide was determined to be 53.8 Å by Guinier analysis of SAXS. The molecular mass of a sphere with an Rg of 53.8 Å is calculated to be 1140 kDa, which is far greater than the observed molecular mass for the 114 kDa chromopeptide. The discrepancy comes from the deviation of the molecular shape from a sphere and/or the dense distribution of electrons in the marginal area of the molecule. The apparent molecular mass of the chromopeptide was determined to be 314-318 kDa from steric exclusion column chromatography, indicating a nonglobular molecular shape. Assuming the overall shape is an ellipsoid of revolution, a prolate one (a = 24, b = 115 A ̊ and an oblate one (a = 85, b = 9 A ̊), where a and b are the equatorial diameter and the half-axial length, respectively, are calculated to have an Rg of 53.8 Å and a molecular mass of 228 kDa. The Rg of the 121 kDa phytochrome was 54.0 Å, suggesting that the terminal polypeptide(s), which is iacking in the 114 kDa chromopeptide, contributes only slightly to the molecular dimension. The apparent molecular mass of the chromopeptide increased to 321-324 kDa after red-light irradiation, indicating that the far-red-light-absorbing form (Pfr) of the chromopeptide has a different molecular dimension from that of Pr. The Guinier analysis of the SAXS of the chromopeptide in Pfr, however, was not successful, owing to the low content of Pfr and the aggregation after saturating red-light irradiation.

Original languageEnglish
Pages (from-to)297-305
Number of pages9
JournalBiochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
Volume953
Issue numberC
DOIs
Publication statusPublished - 1988
Externally publishedYes

Fingerprint

Phytochrome
Peas
Molecular mass
X ray scattering
X-Rays
Light
Gel Chromatography
Column chromatography
Phosmet
Edetic Acid
Irradiation
Electrons
Dimers
Peptides
Agglomeration
Molecules

Keywords

  • Conformational change
  • Photoreceptor protein
  • Phytochrome
  • Protein conformation
  • Radius of gyration
  • Small-angle X-ray scattering

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology
  • Structural Biology

Cite this

Small-angle X-ray scattering studies on the macromolecular structure of the red-light-absorbing form of 121 kDa pea phytochrome and its 114 kDa chromopeptide. / Tokutomi, Satoru; Kataoka, Mikio; Sakai, Jun; Nakasako, Masayoshi; Tokunaga, Fumio; Tasumi, Mitsuo; Furuya, Masaki.

In: Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular, Vol. 953, No. C, 1988, p. 297-305.

Research output: Contribution to journalArticle

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abstract = "The macromolecular structure of 121 kDa pea phytochrome and its 114 kDa chromopeptide was studied by small-angle X-ray scattering (SAXS) and steric exclusion column chromatography. The molecular mass of the 114 kDa chromopeptide in the red-light-absorbing form (Pr) was determined to be 228 kDa in 100 mM potassium phosphate and 1 mM EDTA (pH 7.8) by SAXS, indicating that the chromopeptide is a dimer. The radius of gyration (Rg) of the chromopeptide was determined to be 53.8 {\AA} by Guinier analysis of SAXS. The molecular mass of a sphere with an Rg of 53.8 {\AA} is calculated to be 1140 kDa, which is far greater than the observed molecular mass for the 114 kDa chromopeptide. The discrepancy comes from the deviation of the molecular shape from a sphere and/or the dense distribution of electrons in the marginal area of the molecule. The apparent molecular mass of the chromopeptide was determined to be 314-318 kDa from steric exclusion column chromatography, indicating a nonglobular molecular shape. Assuming the overall shape is an ellipsoid of revolution, a prolate one (a = 24, b = 115 A ̊ and an oblate one (a = 85, b = 9 A ̊), where a and b are the equatorial diameter and the half-axial length, respectively, are calculated to have an Rg of 53.8 {\AA} and a molecular mass of 228 kDa. The Rg of the 121 kDa phytochrome was 54.0 {\AA}, suggesting that the terminal polypeptide(s), which is iacking in the 114 kDa chromopeptide, contributes only slightly to the molecular dimension. The apparent molecular mass of the chromopeptide increased to 321-324 kDa after red-light irradiation, indicating that the far-red-light-absorbing form (Pfr) of the chromopeptide has a different molecular dimension from that of Pr. The Guinier analysis of the SAXS of the chromopeptide in Pfr, however, was not successful, owing to the low content of Pfr and the aggregation after saturating red-light irradiation.",
keywords = "Conformational change, Photoreceptor protein, Phytochrome, Protein conformation, Radius of gyration, Small-angle X-ray scattering",
author = "Satoru Tokutomi and Mikio Kataoka and Jun Sakai and Masayoshi Nakasako and Fumio Tokunaga and Mitsuo Tasumi and Masaki Furuya",
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T1 - Small-angle X-ray scattering studies on the macromolecular structure of the red-light-absorbing form of 121 kDa pea phytochrome and its 114 kDa chromopeptide

AU - Tokutomi, Satoru

AU - Kataoka, Mikio

AU - Sakai, Jun

AU - Nakasako, Masayoshi

AU - Tokunaga, Fumio

AU - Tasumi, Mitsuo

AU - Furuya, Masaki

PY - 1988

Y1 - 1988

N2 - The macromolecular structure of 121 kDa pea phytochrome and its 114 kDa chromopeptide was studied by small-angle X-ray scattering (SAXS) and steric exclusion column chromatography. The molecular mass of the 114 kDa chromopeptide in the red-light-absorbing form (Pr) was determined to be 228 kDa in 100 mM potassium phosphate and 1 mM EDTA (pH 7.8) by SAXS, indicating that the chromopeptide is a dimer. The radius of gyration (Rg) of the chromopeptide was determined to be 53.8 Å by Guinier analysis of SAXS. The molecular mass of a sphere with an Rg of 53.8 Å is calculated to be 1140 kDa, which is far greater than the observed molecular mass for the 114 kDa chromopeptide. The discrepancy comes from the deviation of the molecular shape from a sphere and/or the dense distribution of electrons in the marginal area of the molecule. The apparent molecular mass of the chromopeptide was determined to be 314-318 kDa from steric exclusion column chromatography, indicating a nonglobular molecular shape. Assuming the overall shape is an ellipsoid of revolution, a prolate one (a = 24, b = 115 A ̊ and an oblate one (a = 85, b = 9 A ̊), where a and b are the equatorial diameter and the half-axial length, respectively, are calculated to have an Rg of 53.8 Å and a molecular mass of 228 kDa. The Rg of the 121 kDa phytochrome was 54.0 Å, suggesting that the terminal polypeptide(s), which is iacking in the 114 kDa chromopeptide, contributes only slightly to the molecular dimension. The apparent molecular mass of the chromopeptide increased to 321-324 kDa after red-light irradiation, indicating that the far-red-light-absorbing form (Pfr) of the chromopeptide has a different molecular dimension from that of Pr. The Guinier analysis of the SAXS of the chromopeptide in Pfr, however, was not successful, owing to the low content of Pfr and the aggregation after saturating red-light irradiation.

AB - The macromolecular structure of 121 kDa pea phytochrome and its 114 kDa chromopeptide was studied by small-angle X-ray scattering (SAXS) and steric exclusion column chromatography. The molecular mass of the 114 kDa chromopeptide in the red-light-absorbing form (Pr) was determined to be 228 kDa in 100 mM potassium phosphate and 1 mM EDTA (pH 7.8) by SAXS, indicating that the chromopeptide is a dimer. The radius of gyration (Rg) of the chromopeptide was determined to be 53.8 Å by Guinier analysis of SAXS. The molecular mass of a sphere with an Rg of 53.8 Å is calculated to be 1140 kDa, which is far greater than the observed molecular mass for the 114 kDa chromopeptide. The discrepancy comes from the deviation of the molecular shape from a sphere and/or the dense distribution of electrons in the marginal area of the molecule. The apparent molecular mass of the chromopeptide was determined to be 314-318 kDa from steric exclusion column chromatography, indicating a nonglobular molecular shape. Assuming the overall shape is an ellipsoid of revolution, a prolate one (a = 24, b = 115 A ̊ and an oblate one (a = 85, b = 9 A ̊), where a and b are the equatorial diameter and the half-axial length, respectively, are calculated to have an Rg of 53.8 Å and a molecular mass of 228 kDa. The Rg of the 121 kDa phytochrome was 54.0 Å, suggesting that the terminal polypeptide(s), which is iacking in the 114 kDa chromopeptide, contributes only slightly to the molecular dimension. The apparent molecular mass of the chromopeptide increased to 321-324 kDa after red-light irradiation, indicating that the far-red-light-absorbing form (Pfr) of the chromopeptide has a different molecular dimension from that of Pr. The Guinier analysis of the SAXS of the chromopeptide in Pfr, however, was not successful, owing to the low content of Pfr and the aggregation after saturating red-light irradiation.

KW - Conformational change

KW - Photoreceptor protein

KW - Phytochrome

KW - Protein conformation

KW - Radius of gyration

KW - Small-angle X-ray scattering

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