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Participation of the Surface Structure of Pharaonis Phoborhodopsin, p pR and its A149S and A149V mutants, Consisting of the C‐terminal α‐helix and E–F Loop, in the Complex‐formation with the Cognate Transducer p HtrII, as Revealed by Site‐directed 13 C Solid‐state NMR †
Author(s) -
Kawamura Izuru,
Ikeda Yoichi,
Sudo Yuki,
Iwamoto Masayuki,
Shimono Kazumi,
Yamaguchi Satoru,
Tuzi Satoru,
Saitô Hazime,
Kamo Naoki,
Naito Akira
Publication year - 2007
Publication title -
photochemistry and photobiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.818
H-Index - 131
eISSN - 1751-1097
pISSN - 0031-8655
DOI - 10.1562/2006-06-20-ra-940
Subject(s) - chemistry , helix (gastropod) , crystallography , nmr spectra database , carbon 13 nmr , nuclear magnetic resonance spectroscopy , stereochemistry , spectral line , biology , ecology , physics , astronomy , snail
We have recorded 13 C solid state NMR spectra of [3‐ 13 C]Ala‐labeled pharaonis phoborhodopsin ( p pR) and its mutants, A149S and A149V, complexed with the cognate transducer pharaonis halobacterial transducer II protein ( p HtrII) (1–159), to gain insight into a possible role of their cytoplasmic surface structure including the C‐terminal α‐helix and E–F loop for stabilization of the 2:2 complex, by both cross‐polarization magic angle spinning (CP‐MAS) and dipolar decoupled (DD)‐MAS NMR techniques. We found that 13 C CP‐MAS NMR spectra of [3‐ 13 C]Ala‐ p pR, A149S and A149V complexed with the transducer p HtrII are very similar, reflecting their conformation and dynamics changes caused by mutual interactions through the transmembrane α‐helical surfaces. In contrast, their DD‐MAS NMR spectral features are quite different between [3‐ 13 C]Ala‐ A149S and A149V in the complexes with p HtrII: 13 C DD‐MAS NMR spectrum of [3‐ 13 C]Ala‐A149S complex is rather similar to that of the uncomplexed form, while the corresponding spectral feature of A149V complex is similar to that of p pR complex in the C‐terminal tip region. This is because more flexible surface structure detected by the DD‐MAS NMR spectra are more directly influenced by the dynamics changes than the CP‐MAS NMR. It turned out, therefore, that an altered surface structure of A149S resulted in destabilized complex as viewed from the 13 C NMR spectrum of the surface areas, probably because of modified conformation at the corner of the helix E in addition to the change of hydropathy. It is, therefore, concluded that the surface structure of p pR including the C‐terminal α‐helix and the E–F loops is directly involved in the stabilization of the complex through conformational stability of the helix E.