Photoacoustic Studies of Carboxymyoglobin

Pulsed-laser photoacoustics offers a means for measuring the Fe-CO bond strength in carboxymyoglobin, as well as the dynamics of the protein following cleavage of the Fe-CO bond. Two groups have studied the problem in the past, with conflicting results [Leung et al., Chem. Phys. Letts. 141, 220 (1987); Westrick et al., Biochemistry 26, 8313 (1987)]. Preliminary results from our laboratory support a positve value for the enthalpy of formation of the Fe:CO geminate pair, formed within 10 nsec of photolysis of carboxymyoglobin. Data will be presented on the Fe-CO bond energy, and the volumetric and enthalpic changes which occur in myoglobin following Fe-CO bond cleavage

Photoacoustic Determination of Fluorescent Quantum Yields of Protein Probes

Pulsed-laser photoacoustic methods may be used to determine the fluorescent quantum yields of fluorophores in solution. Of interest to biological spectroscopists are the fluorescent quantum yields of probes bound covalently or noncovalently to proteins. Previous studies (J.R. Small et al., Fluorescence Detection III, E.R. Menzel, ed., SPIE Proceedings 1054, pp. 26-35, 1989) have been extended to examine some common protein probes and their fluorescent quantum yields. Examples include the probes Prodan [6-propionyl-2-(dimethylamino)naphthalene] and Acrylodan [6-acryloyl-2-(dimethylamino)naphthalene] in a variety of protein and solvent environments. We have found that, at the simplest level, the pulsed-laser photoacoustic technique gives us excellent results for the fluorescent quantum yields of fluorophores free in solution, but interestingly anomalous results for the fluorophores bound to proteins. The source of the anomalous protein results has not yet been determined, but several possibilities are discussed

Effects of Solvent Viscosity on the Microsecond Protein Motions of Myoglobin Determined by Pulsed-Laser Photoacoustics

Pulsed-laser photoacoustics offers a means for measuring the Fe-CO bond strength in carboxymyoglobin, as well as the dynamics of the protein following cleavage of the Fe-CO bond. The technique is sensitive to processes which occur on the nsec and psec time scales. Preliminary results from our laboratory support a positive value for the enthalpy of formation of the Fe:CO geminate pair, formed within 10 nsec of photolysis of carboxymyoglobin, with little volumetric change in the protein. A subsequent decay occurs, with a lifetime of about 0.8 gsec at 23 °C, resulting in a volume increase and enthalpic change of the protein. This microsecond decay is temperature dependent and appears to be influenced by solvent viscosity. We are interested in this decay as a relaxation process in myoglobin not generally observable through the spectroscopy of the heme. We will present data on the effect of added glycerol or methanol, the source of the myoglobin, the method of deoxygenation, and the intensity of the photolysis laser pulse on the recovered photoacoustic decay parameters