ABSTRACT
Light-adapted purple membranes were modified with tetranitromethane by a new light-dependent procedure at pH 5.5 which results in a blue-shifted chromophore absorbing at 530nm. This modification affects two aromatic residues. The modified bacteriorhodopsin's ground state chromophore structure is probed by circular dichroism and resonance raman spectroscopy while its photocycle is studied by laser-flash photolysis in the picosecond, microsecond and millisecond time scale. After nitration, the main findings are 1) Interactions between neighboring chromophores are lost, 2) Modified bacteriorhodopsin contains a conformationally changed chromophore but retains a protonated Schiff's base as evidenced by a resonance raman band at 1652 cm-1, 3) A red-shifted intermediate is formed in less than 10 ps after laser excitation, 4) The decay of the M-intermediate is not significantly affected whereas the rise time of the intermediate is enhanced about two fold. These observations are relevant to the role of aromatic acid residues of the apoprotein in the determination of the chromophoric characteristics in bacteriorhodopsin.
Subject(s)
Bacteriorhodopsins/metabolism , Carotenoids/metabolism , Methane/analogs & derivatives , Tetranitromethane/pharmacology , Circular Dichroism , Halobacterium/metabolism , Kinetics , Light , Protein Conformation , SpectrophotometryABSTRACT
Technological advances in picosecond spectroscopy have permitted the mechanisms of various chemical, physical and biological processes to be elucidated and understood to a greater degree than ever before. By means of picosecond emission, absorption and Raman spectroscopy, one can probe and measure directly the transient intermediates and kinetics of primary events in complex biological processes. A description of two current types of laser systems--solid-state and synchronously pumped dye lasers--and their application to determining the primary events in the biological processes of dissociation of oxy- and carboxymyoglobin, excited-state relaxation of porphyrins and visual transduction, illustrate the power of picosecond spectroscopy.
Subject(s)
Myoglobin , Retinal Pigments , Rhodopsin , Spectrum Analysis/methods , Carbon Monoxide , Isomerism , Lasers , Oxygen , Porphyrins , Spectrum Analysis, Raman/methods , Time FactorsABSTRACT
The thermal rearrangements of the bicyclo[2.1.0]pentane-5,2'-methylenecyclopropanes fall into two classes. The first occurs near 80 degrees C and consists of a double epimerization ("bridge flip") which is initiated by cleavage of the bridge bond. An alternative mechanism by way of a trimethylenemethane intermediate is ruled out by an isotopic position-marking experiment. The second rearrangement begins to be detected above 120 degrees C. It gives the isomeric 6- and 7-methylenebicyclo[3.2.0]hept-1-enes. Two possible mechanisms can operate in this complex change, but a choice between them is not yet possible.
