Vectorially oriented fixation of membrane-embedded bacteriorhodopsin onto an inert base

Yoshinori Harada, Kenji Yasuda, Sayuri Nomura, Naoko Kajimura, Yuji C. Sasaki

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

This paper describes a novel surface processing technique aimed at the chemical fixation of proteins on substrate surfaces. The essential feature of this newly developed technique is a combination of chemical and enzymatic processes. A simple technique using chemical modification and selective enzymatic digestion for immobilizing biomembrane-embedded proteins on inorganic solid bases, while strictly regulating their vectorial orientation, was developed. These processes are applicable to a wide range of membrane-embedded and individual proteins, because they exploit the most fundamental principle of proteins, that any protein has at least one N-terminus and one C-terminus. After thorough protection of the carboxyl and amino groups on the molecular surface of bacteriorhodopsin (bR) embedded in purple membrane (PM), in which the bR molecules are uniformly oriented, the derivatized bR was subjected to successive enzymatic digestions to regenerate the unique N-terminal amino group on the molecular surface. The derivatized bR was anchored on an inorganic base by the regenerated amino groups and formed an oriented layered structure. This was proved by analyzing the distance from the base to the gold clusters marking the enzymatically exposed C-termini, which was 4.4 nm, as measured by the fluorescent X-ray interference pattern. This thickness coincides well with that of native PM with embedded bR (4 nm). From the viewpoint of its simplicity, this immobilization process might have an advantage over the former multistep processes for surface functionalization and those for the regulation of the molecular orientation of proteins.

Original languageEnglish
Pages (from-to)1829-1835
Number of pages7
JournalLangmuir
Volume14
Issue number7
DOIs
Publication statusPublished - 1998 Mar 31
Externally publishedYes

ASJC Scopus subject areas

  • Materials Science(all)
  • Condensed Matter Physics
  • Surfaces and Interfaces
  • Spectroscopy
  • Electrochemistry

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