Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2008 Jun 1;64(Pt 6):533-6.
doi: 10.1107/S1744309108013602. Epub 2008 May 23.

Expression, crystallization and preliminary X-ray analysis of an outer membrane protein from Thermus thermophilus HB27

Alexander Brosig  1 , Jutta Nesper, Wolfram Welte, Kay Diederichs
Affiliations

Expression, crystallization and preliminary X-ray analysis of an outer membrane protein from Thermus thermophilus HB27

Alexander Brosig et al. Acta Crystallogr Sect F Struct Biol Cryst Commun. .

Abstract

The cell envelope of the thermophilic bacterium Thermus thermophilus is multilayered and includes an outer membrane with integral outer membrane proteins that are not well characterized. The hypothetical protein TTC0834 from T. thermophilus HB27 was identified as a 22 kDa outer membrane protein containing eight predicted beta-strands. TTC0834 was expressed with an N-terminal His tag in T. thermophilus HB8 and detected in the S-layer/outer membrane envelope fraction. His-TTC0834 was purified and crystallized under various conditions. Native data sets were collected to 3.2 A resolution and the best diffracting crystals belonged to space group P3(1)21 or P3(2)21, with unit-cell parameters a = b = 166.67, c = 97.53 A.

PubMed Disclaimer

Figures

Figure 1
Figure 1
(a) Expression and cell-envelope localization of His-TTC0834 in T. thermophilus HB8. A Coomassie-stained 12% polyacrylamide gel is shown. Lanes 1 and 2, whole cell extracts of HB8 pMK18 (lane 1) and HB8 pMK18-HisTTC0834 (lane 2). Lanes 3 and 4, cell envelopes of HB8 pMK18 (lane 3) and HB8 pMK18-HisTTC0834 (lane 4). Lane M contains molecular-weight markers (kDa). (b) Fractionation of cell envelopes from HB8 expressing His-TTC0834 by centrifugation in sucrose-density gradients. A Coomassie-stained 12% polyacrylamide gel containing samples of fractions 1–8 (from the top to the bottom of the gradient) is shown. Lane M contains molecular-weight markers (kDa). (c) Heat stability of His-TTC0834. A Coomassie-stained 12% polyacrylamide gel with purified His-TTC0834, loaded either directly (295 K) or after boiling for 15 min (373 K), is shown. Note that His-TTC0834 could not be denatured completely by boiling for 15 min.
Figure 2
Figure 2
(a) Native His-TTC0834 crystals grown in 44% MPD, 0.2 M sodium malonate. (b) Coomassie-stained PAA gel loaded with crystals dissolved in SDS sample buffer. The sample was boiled for 7 min prior to loading.
Figure 3
Figure 3
Diffraction pattern of a His-TTC0834 crystal as shown in Fig. 2 ▶ recorded on beamline ID23-1 at the ESRF.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Albrecht, R., Zeth, K., Söding, J., Lupas, A. & Linke, D. (2006). Acta Cryst. F62, 415–418. - PMC - PubMed
    1. Baetens, M., Legrain, C., Boyen, A. & Glansdorff, N. (1998). Microbiology, 144, 479–492. - PubMed
    1. Cava, F., de Pedro, M. A., Schwarz, H., Henne, A. & Berenguer, J. (2004). Mol. Microbiol.52, 677–690. - PubMed
    1. Diederichs, K. & Karplus, P. A. (1997). Nature Struct. Biol.4, 269–275. - PubMed
    1. Gupta, R. S. (2000). Crit. Rev. Microbiol.26, 111–131. - PubMed

MeSH terms

Substances