Time-Resolved FT-IR Difference Spectroscopy: A Tool to Monitor Molecular Reaction Mechanisms of Proteins

2000 ◽  
pp. 209-250
Keyword(s):  
Reaction Mechanisms ◽  
Difference Spectroscopy ◽  
Time Resolved ◽  
Molecular Reaction ◽  
Ft Ir

Molecular Reaction Mechanisms of Proteins Monitored by Nanosecond Step-Scan FT-IR Difference Spectroscopy

1997 ◽  
Vol 51 (4) ◽  
pp. 558-562 ◽  
Author(s):  
Robin Rammelsberg ◽  
Benedikt Heβling ◽  
Harald Chorongiewski ◽  
Klaus Gerwert
Keyword(s):  
Membrane Protein ◽  
Proton Pump ◽  
Reaction Mechanisms ◽  
Three Dimensional ◽  
Difference Spectroscopy ◽  
Dimensional Representation ◽  
Three Dimensional Representation ◽  
Molecular Reaction ◽  
Ft Ir ◽  
Absorbance Changes

Step-scan FT-IR difference spectroscopy with 100-ns time resolution is applied to the membrane protein bacteriorhodopsin. The experimetal setup and data acquisition are described in great detail. The IR absorbance changes accompanying bacteriorhodopsin's photocycle are presented in a three-dimensional representation. The data quality allows one, for example, to resolve the biphasic rise of the photocycle-intermediate M. A bacteriorhodopsin's proton pump model based on the FT-IR works is presented.

scholarly journals Time-resolved flow-flash FT-IR difference spectroscopy: the kinetics of CO photodissociation from myoglobin revisited

2009 ◽  
Vol 394 (8) ◽  
pp. 2277-2277
Author(s):  
Michael Schleeger ◽  
Christoph Wagner ◽  
Michiel J. Vellekoop ◽  
Bernhard Lendl ◽  
Joachim Heberle
Keyword(s):  
Difference Spectroscopy ◽  
Time Resolved ◽  
Ft Ir ◽  
Kinetics Of

scholarly journals Time-resolved flow-flash FT-IR difference spectroscopy: the kinetics of CO photodissociation from myoglobin revisited

2009 ◽  
Vol 394 (7) ◽  
pp. 1869-1877 ◽  
Author(s):  
Michael Schleeger ◽  
Christoph Wagner ◽  
Michiel J. Vellekoop ◽  
Bernhard Lendl ◽  
Joachim Heberle
Keyword(s):  
Difference Spectroscopy ◽  
Time Resolved ◽  
Ft Ir ◽  
Kinetics Of

Atr/Ft-Ir Difference Spectroscopy of Biological Matter with Microsecond Time Resolution

1996 ◽  
Vol 50 (5) ◽  
pp. 588-596 ◽  
Author(s):  
Joachim Heberle ◽  
Christian Zscherp
Keyword(s):  
Attenuated Total Reflection ◽  
High Temporal Resolution ◽  
Difference Spectroscopy ◽  
Difference Spectra ◽  
Time Resolved ◽  
Precise Control ◽  
Biological Matter ◽  
Vis Spectroscopy ◽  
Ft Ir ◽  
Atr Spectroscopy

Attenuated total reflection (ATR) spectroscopy allows precise control of external parameters vital for proper functioning of biological matter. For the first time in biospectroscopy, ATR difference spectroscopy has been combined with the step-scan technique. The current setup integrates a broad frequency range (800–25,000 cm−1) with high temporal resolution (5 μs). Vibrations are detected that arise from single amino acids (<10−3 absorbance units) of the chromoprotein bacteriorhodopsin. Time-resolved ATR/FT-IR difference spectra are compared with conventional transmission spectra. The high mirror stability enables time-resolved FT-vis spectroscopy of the same sample with the same instrument. Potential applications even to non-light-absorbing biomaterial are discussed.

scholarly journals Partial Reactions of the Na,K-ATPase: Determination of Activation Energies and an Approach to Mechanism

2020 ◽  
Vol 253 (6) ◽  
pp. 631-645
Author(s):  
Hans-Jürgen Apell ◽  
Milena Roudna
Keyword(s):  
Reaction Mechanisms ◽  
Fluorescent Dyes ◽  
Activation Energies ◽  
Mechanistic Models ◽  
Time Resolved ◽  
Conformational Relaxation ◽  
Molecular Reaction ◽  
Rate Limiting ◽  
Induced Reaction

Abstract Kinetic experiments were performed with preparations of kidney Na,K-ATPase in isolated membrane fragments or reconstituted in vesicles to obtain information of the activation energies under turnover conditions and for selected partial reactions of the Post-Albers pump cycle. The ion transport activities were detected with potential or conformation sensitive fluorescent dyes in steady-state or time-resolved experiments. The activation energies were derived from Arrhenius plots of measurements in the temperature range between 5 °C and 37 °C. The results were used to elaborate indications of the respective underlying rate-limiting reaction steps and allowed conclusions to be drawn about possible molecular reaction mechanisms. The observed consequent alteration between ligand-induced reaction and conformational relaxation steps when the Na,K-ATPase performs the pump cycle, together with constraints set by thermodynamic principles, provided restrictions which have to be met when mechanistic models are proposed. A model meeting such requirements is presented for discussion. Graphic Abstract

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