MbCO recombination

FIGURE 1.13 Experimental results (points) obtained from pump-probe (left) and transient phase grating (right) measurements of MbCO recombination as a function of time, t, following photodissociation. Solid lines were computed from equations (26) and (27), and the best-fit parameter values are listed in Table 1.2. (From Walther, M., Raicu, V., Ogilvie, J. R, Phillips, R., Kluger, R., and Miller, R. J. D. 2005. J. Phys. Chem. B 109 20605-11. With permission.)... 

Fig. 4. Difference changes in the Soret absorbance region with recombination at 40 K. If the MbCO recombined with no variation in rate with spectral position, then the plot should be flat. The two double derivative peaks are due to Mb (deoxy) on the right-hand side and MbCO on the left. The dashed line shows a spectrum of partially photolyzed MbCO to show the position of the Mb and MbCO peaks. (From Ormos et at )...

Nanosecond time-resolved crystallography of MbCO has been discussed in Section 3.7.2.3 of Chapter 3.46 After firing a 10-ns burst of laser light to break the CO-Fe bond, these researchers produced a diffraction image of the crystal through application of a 150-ps X-ray pulse. They are able to show release of the CO molecule, displacement of the Fe ion toward the proximal histidine, and recombination of the dissociated CO by about 100 ps. Essentially their results compare well with other spectroscopic studies of HbCO, MbCO and their models. 

Pathway I was observed for all the 02 complexes studied, strained or unstrained, as well as for the unstrained CO-complexes. This particular pathway is the same one observed in the photodissociation of the natural heme complexes (3,4) (HbCO, MbCO, HbO and MbO ) with the exception that there is no detectable geminate recombination to the limit of our experiment, 50 ps. Pathway II, observed for the strained-CO complexes, reveals the presence of a fifth intermediate X found early in the dissociation that is either absent or undetectable in the natural or synthetic heme complexes following pathway I. The kinetics associated with the evolution of these intermediates will be discussed shortly. First, it is appropriate to examine in some detail the experimental AA difference spectra of two representative complexes, 1 -CO and 1-ET-CO. A discussion of 1-ST-CO and l-ET-O is also included for comparative purposes. ... 

Chance and co-workers have designed a flow system where the protein is continuously pumped optically using a tungsten or xenon flash lamp (764 nm). Using continuous illumination for various times and temperatures. Chance et al. have observed three intermediate states upon MbCO photolysis. At 40 K, a state with a recombination rate constant of 2 x 10 /s has been identified from two slower states with rate constants of 10 /s. 

Many methods of investigation of protein-ligand binding kinetics that are based on linear processes are of a pump-probe type. In this approach an optical pulse, called a pump, starts a photoreaction (such as dissociation of MbCO into Mb and CO), and its progress is probed a time At later. The probe could be, for example, a weak laser pulse, which detects the spectral changes in the heme during the protein-ligand recombination, or an x-ray pulse, which allows determination of the protein structure at a particular instant in time. 

Figure 9 Geminate recombination after photolysis of MbCO ( ) and microperoxidase-CO ( ). The survival fraction refers to the population that remains unbound after photolysis. The population was determined by measuring the IR absorbance at frequencies corresponding to the peak of the bound CO stretch. (Adapted from Ref. 67.)...

An example of the process described by Eq. (56) is given by the low-temperature (below 200 K) recombination kinetics CO to myoglobin (Mb, a heme group with a central iron atom) after photolysis of carboxymyoglobin MbCO [40,41], A short account of the kinetics which have been described by Eq. (56) is given in Ref. 37. 

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