Purification of the photosynthetic reaction center from Heliobacterium modesticaldum.

Iosifina Sarrou, Zahid Khan, John Cowgill, S. Lin, Daniel Brune, Steven Romberger, John H. Golbeck, Kevin E. Redding

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

We have developed a purification protocol for photoactive reaction centers (HbRC) from Heliobacterium modesticaldum. HbRCs were purified from solubilized membranes in two sequential chromatographic steps, resulting in the isolation of a fraction containing a single polypeptide, which was identified as PshA by LC-MS/MS of tryptic peptides. All polypeptides reported earlier as unknown proteins (in Heinnickel et al., Biochemistry 45:6756-6764, 2006; Romberger et al., Photosynth Res 104:293-303, 2010) are now identified by mass spectrometry to be the membrane-bound cytochrome c (553) and four different ABC-type transporters. The purified PshA homodimer binds the following pigments: 20 bacteriochlorophyll (BChl) g, two BChl g', two 8(1)-OH-Chl a (F), and one 4,4'-diaponeurosporene. It lacks the PshB polypeptide binding the F(A) and F(B) [4Fe-4S] clusters. It is active in charge separation and exhibits a trapping time of 23 ps, as judged by time-resolved fluorescence studies. The charge recombination rate of the P(800) (+)F(X)(-) state is 10-15 ms, as seen before. The purified HbRC core was able to reduce cyanobacterial flavodoxin in the light, exhibiting a K (M) of 10 μM and a k (cat) of 9.5 s(-1) under near-saturating light. There are ~1.6 menaquinones per HbRC in the purified complex. Illumination of frozen HbRC in the presence of dithionite can cause creation of a radical at g = 2.0046, but this is not a semiquinone. Furthermore, we show that high-purity HbRCs are very stable in anoxic conditions and even remain active in the presence of oxygen under low light.

Original languageEnglish (US)
Pages (from-to)291-302
Number of pages12
JournalPhotosynthesis Research
Volume111
Issue number3
DOIs
StatePublished - Jan 1 2012

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Heliobacterium
photosynthetic reaction centers
Photosynthetic Reaction Center Complex Proteins
Purification
polypeptides
Bacteriochlorophylls
Peptides
Light
ABC transporters
Flavodoxin
cytochrome c
menaquinones
Dithionite
Membranes
Vitamin K 2
anaerobic conditions
purity
biochemistry
Biochemistry
ATP-Binding Cassette Transporters

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Plant Science
  • Cell Biology

Cite this

Sarrou, I., Khan, Z., Cowgill, J., Lin, S., Brune, D., Romberger, S., ... Redding, K. E. (2012). Purification of the photosynthetic reaction center from Heliobacterium modesticaldum. Photosynthesis Research, 111(3), 291-302. https://doi.org/10.1007/s11120-012-9726-9
Sarrou, Iosifina ; Khan, Zahid ; Cowgill, John ; Lin, S. ; Brune, Daniel ; Romberger, Steven ; Golbeck, John H. ; Redding, Kevin E. / Purification of the photosynthetic reaction center from Heliobacterium modesticaldum. In: Photosynthesis Research. 2012 ; Vol. 111, No. 3. pp. 291-302.
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Sarrou, I, Khan, Z, Cowgill, J, Lin, S, Brune, D, Romberger, S, Golbeck, JH & Redding, KE 2012, 'Purification of the photosynthetic reaction center from Heliobacterium modesticaldum.', Photosynthesis Research, vol. 111, no. 3, pp. 291-302. https://doi.org/10.1007/s11120-012-9726-9

Purification of the photosynthetic reaction center from Heliobacterium modesticaldum. / Sarrou, Iosifina; Khan, Zahid; Cowgill, John; Lin, S.; Brune, Daniel; Romberger, Steven; Golbeck, John H.; Redding, Kevin E.

In: Photosynthesis Research, Vol. 111, No. 3, 01.01.2012, p. 291-302.

Research output: Contribution to journalArticle

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T1 - Purification of the photosynthetic reaction center from Heliobacterium modesticaldum.

AU - Sarrou, Iosifina

AU - Khan, Zahid

AU - Cowgill, John

AU - Lin, S.

AU - Brune, Daniel

AU - Romberger, Steven

AU - Golbeck, John H.

AU - Redding, Kevin E.

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