The ability of actinic light to modify the bacteriorhodopsin photocycle revisited: heterogeneity vs photocooperativity

Abstract

In 1995, evidence both for photocooperativity and for heterogeneity as possible explanations for the ability of actinic light to modify the kinetics and pathways of the bacteriorhodopsin (BR) photocycle was reviewed ( Shrager, R. I., Hendler, R. W., and Bose, S. (1995) Eur. J. Biochem. 229, 589-595 ). Because both concepts could be successfully modeled to experimental data and there was suggestive published evidence for both, it was concluded that both photocooperativity and heterogeneity may be involved in the adaptation of the BR photocycle to different levels of actinic light. Since that time, more information has become available and it seemed appropriate to revisit the original question. In addition to the traditional models based on all intermediates in strict linear sequences, we have considered both homogeneous and heterogeneous models with branches. It is concluded that an explanation based on heterogeneity is more likely to be the true basis for the variation of the properties of the photocycle caused by changes in actinic light intensity. On the basis of new information presented here, it seems that a heterogeneous branched model is more likely than one with separate linear sequences.

Figure 1

Corrections for light-scattering and offset. The curve labeled CBR in panel A comprises two wavelength anges, one from 391 to 521 nm and the other from 541 to 675 nm. The curve labeled Cbg was constructed, as described in the text, by combining the curve CBR with another spectrum for BR that extended the wavelength to 735 nm and then fitting the low and high wavelength light-scattering regions to a cubic polynomial in order to connect the two light-scattering regions. The corrected curve in panel B was obtained by subtracting Cbg from CBR. The curve labeled BR in panel C is for the BR used in the saturation data displayed in Figure 1. It contains some light-scattering and offset. The corrected curve in panel D was obtained by subtracting Cbg from BR and then for an offset of 0.0096 A. See text for further details.

Figure 2

Experimental laser saturation curves. BR turnover was initiated by laser flashes of increasing intensity as described in Experimental Procedures. The units of the Y-axis scale represent the number of monomers per trimer that cycle during the photo-cycle, as well as the sum of both M_f and M_s per trimer. Also shown are the fractions of each relative to the total M (M_tot).

Figure 3

Ohno’s model, assuming 3 monomers turnover per trimer. Predicted results based on the model of Ohno where p is the probability that a given monomer will be hit by at least one photon. The units of the Y-axis are the same as described in the legend to Figure 1.

Figure 4

Tokaji’s original model (15). The predicted relations as functions of p (probability for a monomer being hit by a photon) are shown for total monomers of BR cycling per trimer, monomers of M_f and M_s cycling per trimer, fraction M_f/M_tot and fraction M_s/M_tot. See text for further details and Figure 2 for definition of Y-axis units.

Figure 5

Hendler’s heterogeneous model (1). Predicted results based on Hendler’s model as described in the text. See legend to Figure 1 for definition of Y-axis units.

Figure 6

Plots of all functions involving M_f and M_s vs p (probability of photon absorption). Panel A. Experimental data. Panel B. Simulated data based on the homogeneous, photocooperative, branched model.

Figure 7

Plots of all functions involving M_fM_s vs laser intensity. Panel A. Experimental data. Panel B. Simulated data based on the homogeneous, photocooperative, branched model.

Figure 8

Two exponential fits to experimental data for M_tot (A) and simultaneous fitting to M_f (B) and M_s. The data are represented by points and the two exponential fits by lines. Residuals are shown relative to a zero line.