Anna B. Druzhko

Photoinduced transformation of wild-type and D96N-mutant 4-keto-bacteriorhodopsin gelatin films

Abstract Spectral and kinelic transformation studies of gelatin films based on 4-keto wild-type bacteriorhodopsin (BR) and 4-keto D96N mutant BR were carried out using absorbance spectroscopy. Spectral heterogeneity in the 400 nm range, assumed to be associated with the M-intermediate state, previously characterized for films of 4-keto wild-type BR, was observed in the 4-keto D96N mutant BR. This heterogeneity is associated to a greater extent with chromophore replacement than with amino acid replacement. The time constants of the M-decay are larger for all kinetic components for 4-keto D96N mutant in the absence of sodium azide. In addition, the contribution of the most long-lived component of the M-state decay is almost twice as large as for the film without sodium azide as it is for a film with sodium azide. This sodium azide effect on the M-state decay kinetics is opposite to that observed in the 4-keto wild-type BR. The comparison of the kinetics and spectral transformations of both pigments suggests that films containing 4-keto D96N mutant hold greater promise as media for information storage and retrieval.

Phototransformation and proton pumping activity of the 14-fluoro bacteriorhodopsin derivatives

The photoinduced behavior and proton pumping characteristics of some bacteriorhodopsin (BR) analogs with fluorinated chromophores (all-trans 14-fluorinated [14-F] retinal and 13-cis 14-F retinal) derived from wild type (WT) and D96N mutant BR were investigated. These analogs were characterized using spectrophotometry and a highly sensitive electrochemical technique. Similar to the white membrane JW2N, the apomembranes WT ET 1000 and D96N form photoactive pigments with the 14-F chromophores. The resulting analogs have a major absorption band at 588 nm. Red-shifted pigment (lambdamax</=680 nm) has been previously observed as a minor component of the major 587-nm pigment in 14-F BR made with white membrane JW2N. A similar red-shifted pigment is formed under yellow light (lambda>500 nm) only in the 14-F analogs derived from WT ET 1000. The measurements of the photoinduced transformation in 14-F WT analogs show that the photocycle of the major pigment occurs simultaneously with the process in the red region and is partially masked by the formation of the red-shifted species. The 14-F D96N samples have a significantly slower and more complicated photoinduced behavior. Electrochemical measurements show that the photoinduced transformation of the red species is not accompanied by proton transport.

OPTICAL AND ELECTRICAL PROPERTIES OF BACTERIORHODOPSIN LANGMUIR-BLODGETT FILMS : II. D96N MUTANT AND ITS 4-KETO AND 9-DEMETHYL RETINAL ANALOGS

Abstract Langmuir-Blodgett films prepared from purple membranes of the D96N mutant and two of its chromophore analogs (4-keto-retinal and 9-demethyl-retinal) were characterized for M-state kinetics and photocurrent production. These studies indicated that unlike the wild-type (WT) and its identical chromophore analogs, the photocurrent transients were unusual, showing proton release at pH 3.0. Several possible explanations for these results are considered.

Measurement of proton release and uptake by analogs of bacteriorhodopsin

Proton release and subsequent uptake by several forms of bacteriorhodopsin (bR), including 4-keto analogs of wild-type (WT) and D96N and D85N mutants as well as the 9-demethylretinal analog of WT and D96N mutants, have been measured using a highly sensitive electrochemical technique. Release and uptake of protons by bR in membrane patches on a tin oxide electrode produce a current transient whose amplitude is proportional to the rate of pH change at the electrode surface. Profiles of proton release by the analogs vs. pH are substantially different from the profiles of the native proteins.

An Azulenic Bacteriorhodopsin Analog Has Photoinduced Activity

Abstract— The photoinduced changes of absorbance as well as proton release and uptake have been observed for an azulenic analog of wild‐type bacteriorhodopsin. The release and uptake of protons have been measured using a highly sensitive electrochemical technique. Bacteriorhodopsin membrane patches on a tin‐oxide electrode produce a transient photocurrent that is negative for proton release and positive for proton uptake. For azulenic bacteriorhodopsin the photocurrent is approximately 20% of the transient observed with native bacteriorhodopsin. The existence of the photoinduced absorbance changes and the transient photocurrent are important results for gaining further insight into the photoinduced function of bacteriorhodopsin.

Optical characteristics of polymer films based on bacteriorhodopsin for irreversible recording of optical information.

Bacteriorhodopsin (BR)‐containing polymer films have been developed in which photoinduced transformations of BR molecules take place during the process of photoinduced hydroxylaminolysis (PHA). The routine simplified scheme of the phototransformation is B↔M412 in the case of polymer films based on nonmodified BR. In the case of polymer films based on BR modified by hydroxylamine (HA), this scheme is changed to where retinal oxime (RO) is a final product of chemical trapping of retinal by the HA molecule. Under certain conditions, the rate of RO to B regeneration is infinitely low. So, the irreversibility of RO formation allows one to use PHA for the preparation of films for irreversible write once recording. Some optical sensitometric characteristics are compared for polymer‐BR films and polymer‐BR–HA films. It has been shown that the photosensitivity level for polymer‐BR–HA films depends on the time after film preparation. A method is offered to increase the photosensitivity of polymer‐BR–HA films and to avoid the photosensitivity decrease with the time after film preparation.

Photochromic polymer films based on a 14-F bacteriorhodopsin derivative

Spectral and kinetic characteristics were measured for polymer (gelatin) films based on 14-F bacteriorhodopsin (BR), both wild-type (WT) and D96N mutant, to study the peculiarities of photo-induced transformation of the samples. It was demonstrated earlier that incorporation of 14-F retinal into the apomembrane of both 14-F WT and 14-F D96N produces pigments with drastically different photo-induced behavior, regarding the formation and decay of a red-shifted species at 660 nm. However, similar fundamental differences between 14-F WT and 14-F D96N, if embedded into gelatin matrix, were not observed. Results are discussed in the frame of the relationship between the kinetic rates of two photo-induced processes that occur in suspensions and gelatin films of corresponding pigments. These experimental results can possibly be explained by the difference in kinetics for gelatin films and water suspensions. The main factor in altering the kinetics is the relative humidity of gelatin samples. Therefore, the effect of relative humidity on performance of 14-F BR gelatin films, both WT and D96N, was studied. A range of humidity has been defined for each pigment, in which spectral and kinetic characteristics were changed in the desired direction. It was shown that 14-F WT gelatin films may offer a technological advantage relative to those based on 14-F D96N.

Nonlinear polarization interaction in bacteriorhodopsin films with anisotropically saturating absorption

The effect of protein and matrix modifications on the photoanisotropic properties is studied for developing the concept of impact upon the main optical properties of the dynamic optical material based on bacteriorhodopsin (BR) both interaction of transmembrane protein--chromophore complex BR with matrix and interaction of protein opsin with chromophore retinal. Also possibility of the application of BR-films for the light polarization modulator is proposed.

Some methods for irreversible write-once recording in 'biochrom' films

Summary form only given. The data-storage capabilities of the photochromic material biochrom, a film based on bacteriorhodopsin (BR) embedded into a polymeric matrix, are described. The material is capable of recording optical information (upon exposure to light) with constant and reversible storage at high spatial resolution (>5*10/sup 3/ lines/mm), photosensitivity up to 10/sup -3/ J/cm/sup 2/, and high cycling (10/sup 4/ cycles). Various types of recording (holographical, polarizable, by bits, etc.) are possible. Image recording is performed by yellow light and its erasure by blue light.<<ETX>>

Application of polymer films based on bacteriorhodopsin and its analogs for low-light-level imaging systems

In recent years polymer films based on bacteriorhodopsin (BR) have attracted a lot of attention in the area of optical imaging systems. The high photosensitivity of these films allows the processing of low-power optical signals (several mW/cm2 CW gas laser irradiation). Spatial resolution does not fall below 5000 lines/mm, photoresponse time is 50 microsecond(s) and images can be recorded and erased over million cycles. Polymer film with BR combine a dynamic recording with optical image processing. The characteristics of anisotropically-saturating nonlinearity of polymer films with BR allow a suppression of the background with greater intensity than usable signal intensity of be performed. Low saturation intensity of the polymer films with BR allows the operation of the polarization of low-intensity signals to be realized. Nonlinear photoresponse of the high photosensitivity BR genetic variant Asp96-Glu is studied in this work too. We hope that the polymer films based on BR and its analogs will find potential use precisely in the medical low- light-level imaging systems.