Site-Directed Isotope Labeling and ATR-FTIR Difference Spectroscopy of Bacteriorhodopsin: The PeptiDe Carbonyl Group of Tyr 185 Is Structurally Active During the bR → N Transition

Cheryl F.C. Ludlam, Sanjay Sonar, Chan Ping Lee, Matthew A Coleman, Judith Herzfeld, Uttam L. RajBhandary, Kenneth J. Rothschild

Research output: Contribution to journalArticlepeer-review

71 Scopus citations

Abstract

The largest secondary structural change occurs in the bacteriorhodopsin (bR) photocycle during the M →-N transition. In this work site-directed isotope labeling (SDIL) and attenuated total reflection Fourier transform infrared (ATR-FTIR) difference spectroscopy were used to investigate this conformational change. L-Tyrosine containing a 13C isotope at the carbonyl carbon was selectively incorporated at Tyr 57, Tyr 147, and Tyr 185 by SDIL. This involves the cell-free expression of bR in the presence of Escherichia coli suppressor tRNATyrCUAaminoacylated with L-[l-13C]Tyr. ATR-FTIR difference spectroscopy reveals that of the 11 tyrosines, only the peptiDe carbonyl group of Tyr 185 undergoes a significant structural change during the bR → N transition. Along with other spectroscopic evidence, this result suggests that the Tyr 185-Pro 186 region of the protein is structurally active and may function as a hinge which facilitates the tilt of the cytoplasmic portion of the F-helix in bacteriorhodopsin during the M → N transition.

Original languageEnglish (US)
Pages (from-to)2-6
Number of pages5
JournalBiochemistry
Volume34
Issue number1
DOIs
StatePublished - Jan 1 1995
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

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