Cytoplasmic surface structure of bacteriorhodopsin consisting of interhelical loops and C-terminal alpha helix, modified by a variety of environmental factors as studied by C-13-NMR

Citation
S. Yamaguchi et al., Cytoplasmic surface structure of bacteriorhodopsin consisting of interhelical loops and C-terminal alpha helix, modified by a variety of environmental factors as studied by C-13-NMR, EUR J BIOCH, 268(8), 2001, pp. 2218-2228
Citations number
55
Categorie Soggetti
Biochemistry & Biophysics
Journal title
EUROPEAN JOURNAL OF BIOCHEMISTRY
ISSN journal
00142956 → ACNP
Volume
268
Issue
8
Year of publication
2001
Pages
2218 - 2228
Database
ISI
SICI code
0014-2956(200104)268:8<2218:CSSOBC>2.0.ZU;2-6
Abstract
We have examined the C-13-NMR spectra of [3-C-13] Ala-labeled bacteriorhodo psin and its mutants by varying a variety of environmental or intrinsic fac tors such as ionic strength, temperature, pH, truncation of the C-terminal a helix, and site-directed mutation at cytoplasmic loops, in order to gain insight into a plausible surface structure arising from the C-terminal alph a helix and loops. It is found that the surface structure can be characteri zed as a complex stabilized by salt bridges or metal-mediated linkages amon g charged side chains. The surface complex in bacteriorhodopsin is most pro nounced under the conditions of 10 mM NaCl at neutral pH but is destabilize d to yield relaxed states when environmental factors are changed to high io nic strength, low pH and higher temperature. These two states were readily distinguished by associated spectral changes, including suppressed (cross p olarization-magic angle spinning NMR) or displaced (upfield) C-13 signals f rom the C-terminal alpha helix, or modified spectral features in the loop r egion. It is also noteworthy that such spectral changes, when going from th e complexed to relaxed states, occur either when the C-terminal alpha helix is deleted or site-directed mutations were introduced at a cytoplasmic loo p. These observations clearly emphasize that organization of the cytoplasmi c surface complex is important in the stabilization of the three-dimensiona l structure at ambient temperature, and subsequently plays an essential rol e in biological functions.