Effect of introducing different carboxylate-containing side chains at position 85 on chromophore formation and proton transport in bacteriorhodopsin

Duncan A. Greenhalgh, Sriram Subramaniam, Ulrike Alexiev, Harald Otto, Maarten P. Heyn, H. Gobind Khorana

Research output: Contribution to journalArticle

Abstract

During the initial stages of the bacteriorhodopsin photocycle, a proton is transferred from the Schiff base to the deprotonated carboxylate of Asp85. Earlier studies have shown that replacement of Asp85 by Asn completely abolishes proton transport activity, whereas extension of the side chain by an additional carbon-carbon bond (Asp85 → Glu) results in a functional proton pump. Here we show that extension of the Asp85 side chain by two additional bond lengths also results in a functional proton pump as long as the terminal group is a carboxylate moiety. These side chains were created by modification of the cysteine residue in the Asp85 → Cys mutant with either iodoacetic acid or iodoacetamide. In vitro chromophore formation studies show that the rate of Schiff base protonation in mutants that contain a carboxylate at residue 85 is invariably faster than in mutants that contain neutral substitutions at this position. We conclude that in bacteriorhodopsin, there is considerable tolerance in the volume of the side chain that can be accommodated at position 85 and that the presence of a carboxylate at residue 85 is important both for proton pumping and for stabilizing the protonated Schiff base.

Original languageEnglish (US)
Pages (from-to)25734-25738
Number of pages5
JournalJournal of Biological Chemistry
Volume267
Issue number36
StatePublished - Dec 25 1992

Fingerprint

Bacteriorhodopsins
Schiff Bases
Chromophores
Protons
Proton Pumps
Carbon
Iodoacetic Acid
Iodoacetamide
Protonation
Bond length
Cysteine
Substitution reactions

ASJC Scopus subject areas

  • Biochemistry

Cite this

Greenhalgh, D. A., Subramaniam, S., Alexiev, U., Otto, H., Heyn, M. P., & Khorana, H. G. (1992). Effect of introducing different carboxylate-containing side chains at position 85 on chromophore formation and proton transport in bacteriorhodopsin. Journal of Biological Chemistry, 267(36), 25734-25738.

Effect of introducing different carboxylate-containing side chains at position 85 on chromophore formation and proton transport in bacteriorhodopsin. / Greenhalgh, Duncan A.; Subramaniam, Sriram; Alexiev, Ulrike; Otto, Harald; Heyn, Maarten P.; Khorana, H. Gobind.

In: Journal of Biological Chemistry, Vol. 267, No. 36, 25.12.1992, p. 25734-25738.

Research output: Contribution to journalArticle

Greenhalgh, DA, Subramaniam, S, Alexiev, U, Otto, H, Heyn, MP & Khorana, HG 1992, 'Effect of introducing different carboxylate-containing side chains at position 85 on chromophore formation and proton transport in bacteriorhodopsin', Journal of Biological Chemistry, vol. 267, no. 36, pp. 25734-25738.
Greenhalgh, Duncan A. ; Subramaniam, Sriram ; Alexiev, Ulrike ; Otto, Harald ; Heyn, Maarten P. ; Khorana, H. Gobind. / Effect of introducing different carboxylate-containing side chains at position 85 on chromophore formation and proton transport in bacteriorhodopsin. In: Journal of Biological Chemistry. 1992 ; Vol. 267, No. 36. pp. 25734-25738.
@article{dafaccc16e8b46a098bf55285154843a,
title = "Effect of introducing different carboxylate-containing side chains at position 85 on chromophore formation and proton transport in bacteriorhodopsin",
abstract = "During the initial stages of the bacteriorhodopsin photocycle, a proton is transferred from the Schiff base to the deprotonated carboxylate of Asp85. Earlier studies have shown that replacement of Asp85 by Asn completely abolishes proton transport activity, whereas extension of the side chain by an additional carbon-carbon bond (Asp85 → Glu) results in a functional proton pump. Here we show that extension of the Asp85 side chain by two additional bond lengths also results in a functional proton pump as long as the terminal group is a carboxylate moiety. These side chains were created by modification of the cysteine residue in the Asp85 → Cys mutant with either iodoacetic acid or iodoacetamide. In vitro chromophore formation studies show that the rate of Schiff base protonation in mutants that contain a carboxylate at residue 85 is invariably faster than in mutants that contain neutral substitutions at this position. We conclude that in bacteriorhodopsin, there is considerable tolerance in the volume of the side chain that can be accommodated at position 85 and that the presence of a carboxylate at residue 85 is important both for proton pumping and for stabilizing the protonated Schiff base.",
author = "Greenhalgh, {Duncan A.} and Sriram Subramaniam and Ulrike Alexiev and Harald Otto and Heyn, {Maarten P.} and Khorana, {H. Gobind}",
year = "1992",
month = "12",
day = "25",
language = "English (US)",
volume = "267",
pages = "25734--25738",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "36",

}

TY - JOUR

T1 - Effect of introducing different carboxylate-containing side chains at position 85 on chromophore formation and proton transport in bacteriorhodopsin

AU - Greenhalgh, Duncan A.

AU - Subramaniam, Sriram

AU - Alexiev, Ulrike

AU - Otto, Harald

AU - Heyn, Maarten P.

AU - Khorana, H. Gobind

PY - 1992/12/25

Y1 - 1992/12/25

N2 - During the initial stages of the bacteriorhodopsin photocycle, a proton is transferred from the Schiff base to the deprotonated carboxylate of Asp85. Earlier studies have shown that replacement of Asp85 by Asn completely abolishes proton transport activity, whereas extension of the side chain by an additional carbon-carbon bond (Asp85 → Glu) results in a functional proton pump. Here we show that extension of the Asp85 side chain by two additional bond lengths also results in a functional proton pump as long as the terminal group is a carboxylate moiety. These side chains were created by modification of the cysteine residue in the Asp85 → Cys mutant with either iodoacetic acid or iodoacetamide. In vitro chromophore formation studies show that the rate of Schiff base protonation in mutants that contain a carboxylate at residue 85 is invariably faster than in mutants that contain neutral substitutions at this position. We conclude that in bacteriorhodopsin, there is considerable tolerance in the volume of the side chain that can be accommodated at position 85 and that the presence of a carboxylate at residue 85 is important both for proton pumping and for stabilizing the protonated Schiff base.

AB - During the initial stages of the bacteriorhodopsin photocycle, a proton is transferred from the Schiff base to the deprotonated carboxylate of Asp85. Earlier studies have shown that replacement of Asp85 by Asn completely abolishes proton transport activity, whereas extension of the side chain by an additional carbon-carbon bond (Asp85 → Glu) results in a functional proton pump. Here we show that extension of the Asp85 side chain by two additional bond lengths also results in a functional proton pump as long as the terminal group is a carboxylate moiety. These side chains were created by modification of the cysteine residue in the Asp85 → Cys mutant with either iodoacetic acid or iodoacetamide. In vitro chromophore formation studies show that the rate of Schiff base protonation in mutants that contain a carboxylate at residue 85 is invariably faster than in mutants that contain neutral substitutions at this position. We conclude that in bacteriorhodopsin, there is considerable tolerance in the volume of the side chain that can be accommodated at position 85 and that the presence of a carboxylate at residue 85 is important both for proton pumping and for stabilizing the protonated Schiff base.

UR - http://www.scopus.com/inward/record.url?scp=0027051284&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0027051284&partnerID=8YFLogxK

M3 - Article

C2 - 1361187

AN - SCOPUS:0027051284

VL - 267

SP - 25734

EP - 25738

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 36

ER -