Accurate and rapid novel genetic diagnosis for detection of sentinel lymph node metastasis in breast cancer patients.

BACKGROUND: The transcription-reverse transcription concerted reaction (TRC) test is a novel molecular-based procedure, which can assess nodal metastasis accurately and quickly. We examined the usefulness of the TRC test with a double marker, cytokeratin 19 (CK19) and carcinoembryonic antigen (CEA) mRNA, to detect sentinel lymph nodes (SLN) metastasis in breast cancer patients. METHODS: A total of 264 SLNs from 131 breast cancer patients were assigned to a training set (109 SLNs from 50 patients) and validation set (155 SLNs from 81 patients). Cytokeratin 19 and CEA mRNA were detected by TRC tests, and the sensitivity and specificity of the SLN metastasis between the TRC and histology cohorts were compared. RESULTS: Mean copy numbers of CK19 and CEA by TRC tests were increased according to the metastatic size. In the training set, TRC test showed 100% sensitivity, specificity and concordance rates against the permanent histopathology test. In the validation set, sensitivity was 97.1%, specificity was 99.2% and the concordance rate was 99.4%. CONCLUSION: Our results showed that the detection of CK19 and CEA mRNA using the TRC test is, an accurate and rapid method for detection of SLN metastasis and can be applied as an intraoperative molecular diagnosis in breast cancer patients.

Sotrastaurin, a protein kinase C inhibitor, ameliorates ischemia and reperfusion injury in rat orthotopic liver transplantation.

Sotraustaurin (STN), a small molecule, targeted protein kinase C (PKC) inhibitor that prevents T-lymphocyte activation via a calcineurin-independent pathway, is currently being tested in Phase II renal and liver transplantation clinical trials. We have documented the key role of activated T cells in the inflammation cascade leading to liver ischemia/reperfusion injury (IRI). This study explores putative cytoprotective functions of STN in a clinically relevant rat model of hepatic cold ischemia followed by orthotopic liver transplantation (OLT). Livers from Sprague-Dawley rats were stored for 30 h at 4°C in UW solution, and then transplanted to syngeneic recipients. STN treatment of liver donors/recipients or recipients only prolonged OLT survival to >90% (vs. 40% in controls), decreased hepatocellular damage and improved histological features of IRI. STN treatment decreased activation of T cells, and diminished macrophage/neutrophil accumulation in OLTs. These beneficial effects were accompanied by diminished apoptosis, NF-κB/ERK signaling, depressed proapoptotic cleaved caspase-3, yet upregulated antiapoptotic Bcl-2/Bcl-xl and hepatic cell proliferation. In vitro, STN decreased PKCθ/IκBα activation and IL-2/IFN-γ production in ConA-stimulated spleen T cells, and diminished TNF-α/IL-1ß in macrophage-T cell cocultures. This study documents positive effects of STN on liver IRI in OLT rat model that may translate as an additional benefit of STN in clinical liver transplantation.

Evaluation of the SCA instrument for measuring patient satisfaction with cancer care administered via paper or via the Internet.

BACKGROUND: Patients' perspectives provide valuable information on quality of care. This study evaluates the feasibility and validity of Internet administration of Service Satisfaction Scale for Cancer Care (SCA) to assess patient satisfaction with outcome, practitioner manner/skill, information, and waiting/access. PATIENTS AND METHODS: Primary data collected from November 2007 to April 2008. Patients receiving cancer care within 1 year were recruited from oncology, surgery, and radiation clinics at a tertiary care hospital. An Internet-based version of the 16-item SCA was developed. Participants were randomised to Internet SCA followed by paper SCA 2 weeks later or vice versa. Seven-point Likert scale responses were converted to a 0-100 scale (minimum-maximum satisfaction). Response distribution, Cronbach's alpha, and test-retest correlations were calculated. RESULTS: Among 122 consenting participants, 78 responded to initial SCA. Mean satisfaction scores for paper/Internet were 91/90 (outcome), 95/94 (practitioner manner/skill), 89/90 (information), and 86/86 (waiting/access). Response rate and item missingness were similar for Internet and paper. Except for practitioner manner/skill, test-retest correlations were robust r = 0.77 (outcome), 0.74 (information), and 0.75 (waiting/access) (all P < 0.001). CONCLUSIONS: Internet SCA administration is a feasible and a valid measurement of cancer care satisfaction for a wide range of cancer diagnoses, treatment modalities, and clinic settings.

A simple and sensitive method for detection of Bacillus anthracis by loop-mediated isothermal amplification.

AIMS: To develop a rapid and simple system for detection of Bacillus anthracis using a loop-mediated isothermal amplification (LAMP) method and determine the suitability of LAMP for rapid identification of B. anthracis infection. METHODS AND RESULTS: A specific LAMP assay targeting unique gene sequences in the bacterial chromosome and two virulence plasmids, pXO1 and pXO2, was designed. With this assay, it was possible to detect more than 10 fg of bacterial DNA per reaction and obtain results within 30-40 min under isothermal conditions at 63 degrees C. No cross-reactivity was observed among Bacillus cereus group and other Bacillus species. Furthermore, in tests using blood specimens from mice inoculated intranasally with B. anthracis spores, the sensitivity of the LAMP assay following DNA extraction methods using a Qiagen DNeasy kit or boiling protocol was examined. Samples prepared by both methods showed almost equivalent sensitivities in LAMP assay. The detection limit was 3.6 CFU per test. CONCLUSIONS: The LAMP assay is a simple, rapid and sensitive method for detecting B. anthracis. SIGNIFICANCE AND IMPACT OF THE STUDY: The LAMP assay combined with boiling extraction could be used as a simple diagnostic method for identification of B. anthracis infection.

Case Report of Extensive Isolated Spontaneous Celiac Trunk Dissection After Liver Transplantation.

Arterial dissection is a rare complication after liver transplantation (LT). We report a case of extensive isolated spontaneous celiac trunk dissection (ISCTD) up to the proper hepatic artery, left gastric artery, and splenic artery after living donor liver transplantation. A 48-year-old woman with cryptogenic liver cirrhosis underwent living donor liver transplantation. Intraoperative and postoperative Doppler ultrasound revealed sufficient flow in the hepatic artery, portal vein, and hepatic vein. On postoperative day (POD) 10, Doppler ultrasound showed reduction of hepatic arterial flow. On POD 16, a contrast-enhanced computed tomography scan showed that the ISCTD extended to the proper hepatic artery, left gastric artery, and splenic artery with an entry tear on the proximal side of the celiac trunk. Although the computed tomography scan showed ischemia of a small part of the liver, blood flow to the liver was kept to some extent. Because all false lumens were occluded by thrombi and the liver enzyme levels normalized, we chose conservative therapy with antiplatelet agents. The patient was discharged on POD 53. She remains well without any liver dysfunction after 18 months with reduction in all false lumens and a patent hepatic artery. Several cases of ISCTD have been reported apart from LT, most of which were treated with conservative therapy. We conclude that conservative therapy could be the first choice in ISCTD even after LT.

Glucose transport of Haloferax volcanii requires the Na(+)-electrochemical potential gradient and inhibitors for the mammalian glucose transporter inhibit the transport.

The uptake of glucose and its non-metabolizing analogues by Haloferax volcanii, one of the glucose-utilizing Halobacteria, was examined using intact cells and envelope vesicles. Results obtained were: (1) The transport system is inducible. (2) The uptake requires the gradient of Na(+)-electrochemical potential. (3) Inhibitors for mammalian glucose transport also have an effect on this system, implying that the transporters resemble each other. (4) It is suggested that the mobility of the transporter is regulated by the membrane energization.

Mitochondrial membrane potential estimated with the correction of probe binding.

Lipophilic ions are widely used as the probe for estimation of the membrane potential. It is suggested that the correction of the probe binding to the membrane and/or intracellular constituents is a problem to be solved in order to evaluate the membrane potential accurately. Previously, we proposed a method for the correction of the probe binding (Demura, M., Kamo, N. and Kobatake, Y. (1985) Biochim. Biophys. Acta 820, 207-215). In this paper, the method was applied to the determination of the membrane potential of intact mitochondria. The probes used constitute a homologous series of (Phe)3-P+-(CH2)n-CH3 (n = 0-4) and tetraphenylphosphonium (TPP+). Binding of these probes to de-energized mitochondria followed the Langmuir isotherm. However, values of parameters determined at high (50-800 microM) and low (under 20 microM) probe concentrations were different, suggesting the existence at least two, high- and low-affinity, binding sites. With extrapolation to the 'state of no binding', the membrane potential of intact mitochondria was estimated to be -147 mV (interior-negative) when they were energized by 5 mM succinate in medium consisting of 125 mM KCl, 10 mM MgCl2, 5 mM phosphate, 0.4 mM EDTA and 50 mM Tris-HCl (pH 7.5) at 25 degrees C. Parameters appearing in the equation for the correction of probe binding were determined with the use of this value of the membrane potential. The validity of the equation and the value of the parameters were revealed by the fact that after the correction, all probes used gave approximately the same value under the same conditions. We expanded the method so as to include the langmuir adsorption isotherm. When the modified equation is used, the estimated membrane potentials were less dependent on a probe concentration less than 10 microM.

Binding of lipophilic cations to the liposomal membrane: thermodynamic analysis.

Lipophilic ions are widely used as probes for measuring membrane potentials. Since binding of the probes to the membrane interferes with the accurate estimation of the membrane potential, it is necessary to clarify the characteristics of probe binding to membranes. The present paper deals with the binding of lipophilic cations to liposomes. The results can be summarized as follows: (1) The binding of triphenylmethylphosphonium, its homologues and tetraphenylphosphonium to liposomes of dipalmitoylphosphatidylcholine followed the Langmuir adsorption isotherm. (2) Spin-labeled lipophilic cations were synthesized and the binding to liposomes of egg phosphatidylcholine was examined. The binding also followed the Langmuir adsorption isotherm. The dissociation constant (the concentration giving half-maximal binding), K, was independent of the temperature, indicating that the binding is entropy-driven. (3) The binding was influenced by the fluidity of the membrane. Except in the case of triphenylmethylphosphonium (TPMP+), K and A (maximum amounts of binding) increased above the transition temperature. In other words, above the phase transition temperature the binding affinity is decreased, while maximum amounts of binding are increased for all phosphoniums used except TPMP+.

In archaebacteria, there is a doxorubicin efflux pump similar to mammalian P-glycoprotein.

We selected for study an anthracycline-resistant mutant from the archaebacteria Haloferax volcanii. This resistance was reversed by a Ca(2+)-channel antagonist, nifedipine (NDP). This resistance and its reversal by NDP suggest P-glycoprotein (Pgp) to be responsible for maintaining an anticancer drug concentration below the cytotoxic level. Using rhodamine 123 (RH123) as a substrate for Pgp, we then examined whether the resistance to anthracyclines in this bacteria might involve a Pgp-like anthracycline efflux pump. RH123 accumulation by the bacteria was determined with flow cytometry. A steady-state RH123 accumulation by the resistant cells revealed approx. one-fifteenth of that by the wild-type cells, which could be remarkably enhanced by NDP. The other modulators of Pgp, diltiazem and verapamil, also enhanced RH123 accumulation in resistant cells. The uncoupler FCCP completely restored RH123 accumulation in resistant cells to the wild-type cell level. RH123 unidirectional efflux from resistant cells after its preloading revealed much greater than that from wild-type cells, which was remarkably inhibited by FCCP. These confirmed that RH123 low accumulation involves its active efflux mechanism. Taken together, the present study indicated that lower evolutionary archaebacteria might also express a Pgp-like protein very similar to mammalian Pgp.

Measurement of plasma membrane potential in isolated rat hepatocytes using the lipophilic cation, tetraphenylphosphonium: correction of probe intracellular binding and mitochondrial accumulation.

The lipophilic cation tetraphenylphosphonium (TPP+) has been extensively utilized as the probe for the membrane potential (Vm) in various cells. For application to mammalian cells, however, two serious problems require resolution: (1), correction of TPP+ binding to intracellular constituents and (2), estimation of the considerable TPP+ accumulation in mitochondria. We propose here a simple corrective method for the TPP+ binding and its accumulation. TPP+ distribution is assumed as: (1), two compartments (a cytosolic and a mitochondrial space); (2), a proportional relationship between TPP+ bound amount and its unbound concentration in each compartment. We theoretically derived the simple equation: Vm = - RT/F ln(C/Mphys ratio/C/Mabol ratio) where R, T and F have their usual thermodynamic significance. Here, the C/M ratio is defined as the ratio of TPP+ concentration of apparent intracellular to extracellular space. The suffixes phys and abol, respectively, mean the physiological and solely Vm-abolished conditions. This equation was checked with hepatocytes, because estimating hepatocytes Vm with TPP+ distribution is not considered possible because of the relatively high mitochondrial content. The selective Vm abolition was achieved by permeabilization with 20 microM of amphotericin B. The Vm value was, thus, estimated to be -38.6 +/- 0.3 mV, compatible with those obtained with microelectrodes in other laboratories. Vm in hepatocytes is composed of transmembrane K+ diffusion potential (-20.6 +/- 0.3 mV) and electrogenic Na+/K(+)-ATPase (-19.6 +/- 0.4 mV). Addition of rheogenic L-alanine caused a transient but significant depolarization (from control to -34 +/- 0.3 mV). These results taken together indicate that hepatocyte Vm can be accurately determined with the present simple method, so that it may possibly be applicable to the evaluation of Vm in other mammalian cells.

Pysicochemical studies of taste reception. V. Suppressive effect of salts on sugar response of the frog.

The tast responses of frog to various kinds of sugars were measured quantitatively by use of the glossopharyngeal nerve activity under an appropriate condition where the water response was completely suppressed. The concentration dependences of response of frog tongue to D-fructose, D-glucose, and sucrose were almost the same, D-galactose, however, elicited a much larger response in comparison with the other sugars in the whole range of concentrations examined. The sugar response was suppressed extensively by the presence of small amount of salts in the stimulating sugar solution. The suppressive effects of NaCl, KCl, MgCl2, MgSO4, and K4Fe(CN)6 were examined with a fixed concentration of sugar. The results obtained with these salts, added in various concentrations, fell on a single curve when the data were plotted against the ionic strength in the stimulating solution. The present results were consistent with the notion that the taste receptor potential for salts or acids is attributable to a change in the phase boundary potential at the membrane-solution interface as proposed in the previous papers of this series.

Selective reaction of hydroxylamine with chromophore during the photocycle of pharaonis phoborhodopsin.

Phoborhodopsin (pR; also called sensory rhodopsin II, sRII) is a receptor of negative phototaxis of Halobacterium salinarum, and pharaonis phoborhodopsin (ppR; also pharaonis sensory rhodopsin II, psRII) is a corresponding protein of Natronobacterium pharaonis. These receptors contain retinal as a chromophore which binds to a lysine residue via Schiff base. This Schiff base can be cleaved with hydroxylamine to loose their color (bleaching). In dark, the bleaching rate of ppR was very slow whereas illumination accelerated considerably the bleaching rate. Addition of azide accelerated the decay of the M-intermediate while its formation (decay of the L-intermediate) is not affected. The bleaching rate of ppR under illumination was decreased by addition of azide. Essentially no reactivity with hydroxylamine under illumination was observed in the case of D75N mutant which lacks the M-intermediate in its photocycle. Moreover, we provided illumination by flashes to ppR in the presence of varying concentrations of azide to measure the bleaching rate per one flash. A good correlation was obtained between the rate and the mean residence time, MRT, which was calculated from flash photolysis data of the M-decay. These findings reveal that water-soluble hydroxylamine reacts selectively with the M-intermediate and its implication was discussed.

Selective electrode for dibenzyl dimethyl ammonium cation as indicator of the membrane potential in biological systems.

The electrode sensitive to dibenzyl dimethyl ammonium (DDA+), which is considered to be an indicator of the membrane potential, was constructed by using tetraphenyl borone (TPB-) embedded in dichloroethane. Rapid and Nernstian responses were exhibited against DDA+ solutions ranging between 10(-2) and 3 - 10(-6) M in concentration. High selectivity for DDA+ was observed in the presence of various inorganis salts, ADP, ATP, oxidizable substrates and sugars. The electrode developed here was used to measure the DDA+ uptake in Streptococcus faecalis and the results agreed with those reported by Harold, F.M. and Papineau, D. ((1972) J. Membrane Biol. 8, 27-44 and 45-62). While they determined the DDA+ concentration in the medium by measuring the absorbance of the filtrate treated with the ion-exchangers, the electrode can measure directly the DDA+ concentration in the bacterial suspension without any any pretreatment. It was also shown that the electrode can measure the DDA+ uptake in mitochondria during energization.

Physicochemical studies of taste reception. III. Interpretation of the water response in taste reception.

The model membrane composed of a Millipore filter paper and the total lipids from bovine tongue epithelium or phosphatidylcholine from egg yolk simulated well the water response of a living taste cell, The water response observed with the model membrane adapted to various salt solutions was interpreted in terms of changes in electric potential at the membrane-solution interface, i.e. the water response was attributed to the e.m.f. change produced by diffusion of the electrolytes dissolved in (or adsorbed on) the membrane surface into the bulk solution. The water response of the frog tongue was also investigated by measuring the neural response of the glossopharyngeal nerve. The results obtained were consistent with the mechanism proposed in the present paper. The response of the frog to Ca2+ was examined under the condition where the water response was suppressed, and it was concluded that the water response of the frog is different from the response to Ca2+.

Partitioning of triphenylalkylphosphonium homologues in gel bead-immobilized liposomes: chromatographic measurement of their membrane partition coefficients.

Unilamellar liposomes of small or large size, SUVs and LUVs, respectively, were stably immobilized in the highly hydrophilic Sepharose 4B or Sephacryl S-1000 gel beads as a membrane stationary phase for immobilized liposome chromatography (ILC). Lipophilic cations of triphenylmethylphosphonium and tetraphenylphosphonium (TPP+) have been used as probes of the membrane potential of cells. Interaction of TPP+ and triphenylalkylphosphonium homologues with the immobilized liposomal membranes was shown by their elution profiles on both zonal and frontal ILC. Retardation of the lipophilic cations on the liposome gel bed was increased as the hydrophobicity of the cations increased, indicating the partitioning of lipophilic cations into the hydrocarbon region of the membranes. The cations did not retard on the Sepharose or Sephacryl gel bed without liposomes, confirming that the cations only interact with the immobilized liposomes. Effects of the solute concentration, flow rate, and gel-matrix substance on the ILC were studied. The stationary phase volume of the liposomal membranes was calculated from the volume of a phospholipid molecule and the amount of the immobilized phospholipid, which allowed us to determine the membrane partition coefficient (KLM) for the lipophilic cations distributed between the aqueous mobile and membrane stationary phases. The values of KLM were generally increased with the hydrophobicity of the solutes increased, and were higher for the SUVs than for the LUVs. The ILC method described here can be applied to measure membrane partition coefficients for other lipophilic solutes (e.g., drugs).

Measurement of membrane potential in polymorphonuclear leukocytes and its changes during surface stimulation.

The membrane potential of guinea pig polymorphonuclear leukocytes has been assessed with two indirect probes, tetraphenylphosphonium (TPP+) and 3,3'-dipropylthiadicarbocyanine (disS-C3-(5)). The change in TPP+ concentration in the medium was measured with a TPP+-selective electrode. By monitoring differences in accumulation of TPP+ in media containing low and high potassium concentrations, a resting potential of -58.3 mV was calculated. This potential is composed of a diffusion potential due to the gradient of potassium, established by the Na+, K+ pump, and an electrogenic potential. The chemotactic peptide fMet-Leu-Phe elicits a rapid efflux of accumulated TPP+ (indicative of depolarization) followed by its reaccumulation (indicative of repolarization). In contrast, stimulation with concanavalin A results in a rapid and sustained depolarization without a subsequent repolarization. The results obtained with TPP+ and diS-C3-(5) were comparable. Such changes in membrane potential were observed in the absence of extracellular sodium, indicating that an inward movement of sodium is not responsible for the depolarization. Increasing potassium levels, which lead to membrane depolarization, had no effect on the oxidative metabolism in nonstimulated or in fMet-Leu-Phe-stimulated cells. Therefore, it seems unlikely that membrane depolarization per se is the immediate stimulus for the respiratory burst.

Formation and characterization of planar lipid bilayer membranes from synthetic phytanyl-chained glycolipids.

The formability, current-voltage characteristics and stability of the planar lipid bilayer membranes from the synthetic phytanyl-chained glycolipids, 1, 3-di-O-phytanyl-2-O-(beta-glycosyl)glycerols (Glc(Phyt)(2), Mal(N)(Phyt)(2)) were studied. The single bilayer membranes were successfully formed from the glycolipid bearing a maltotriosyl group (Mal(3)(Phyt)(2)) by the folding method among the synthetic glycolipids examined. The membrane conductance of Mal(3)(Phyt)(2) bilayers in 100 mM KCl solution was significantly lower than that of natural phospholipid, soybean phospholipids (SBPL) bilayers, and comparable to that of 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhPC) bilayers. From the permeation measurements of lipophilic ions through Mal(3)(Phyt)(2) and DPhPC bilayers, it could be presumed that the carbonyl groups in glycerol backbone of the lipid molecule are not necessarily required for the total dipole potential barrier against cations in Mal(3)(Phyt)(2) bilayer. The stability of Mal(3)(Phyt)(2) bilayers against long-term standing and external electric field change was rather high, compared with SBPL bilayers. Furthermore, a preliminary experiment over the functional incorporation of membrane proteins was demonstrated employing the channel proteins derived from octopus retina microvilli vesicles. The channel proteins were functionally incorporated into Mal(3)(Phyt)(2) bilayers in the presence of a negatively charged glycolipid. From these observations, synthetic phytanyl-chained glycolipid bilayers are promising materials for reconstitution and transport studies of membrane proteins.

Environment around the chromophore in pharaonis phoborhodopsin: mutation analysis of the retinal binding site.

Phoborhodopsin (pR or sensory rhodopsin II, sRII) and pharaonis phoborhodopsin (ppR or pharaonis sRII, psRII) have a unique absorption maximum (lambda(max)) compared with three other archaeal rhodopsins: lambda(max) of pR and ppR is approx. 500 nm and of others (e.g. bacteriorhodopsin, bR) is 560-590 nm. To determine the residue contributing to the opsin shift from ppR to bR, we constructed various ppR mutants, in which a single residue was substituted for a residue corresponding to that of bR. The residues mutated were those which differ from that of bR and locate within 5 A from the conjugated polyene chain of the chromophore or any methyl group of the polyene chain. The shifts of lambda(max) of all mutants were small, however. We constructed a mutant in which all residues which differ from those of bR in the retinal binding site were simultaneously substituted for those of bR, but the shift was only from 499 to 509 nm. Next, we constructed a mutant in which 10 residues located within 5 A from the polyene as described above were simultaneously substituted. Only 44% of the opsin shift (lambda(max) of 524 nm) from ppR to bR was obtained even when all amino acids around the chromophore were replaced by the same residues as bR. We therefore conclude that the structural factor is more important in accounting for the difference of lambda(max) between ppR and bR rather than amino acid substitutions. The possible structural factors are discussed.

Gustatory responses of eel palatine receptors to amino acids and carboxylic acids.

The gustatory receptors of the eel palate were found to be extremely sensitive to amino acids and carboxylic acids. The results obtained are as follows: (a) 11 amino acids which are among naturally occurring amino acids elicited responses in the palatine nerve, but 9 amino acids did not elicit a response even at a high concentration. The effect of D-amino acids was always much less than that of their corresponding L-isomers. There was no appreciable difference in the effectiveness of an alpha-amino acid (alpha-alanine) and beta-amino acid (beta-alanine). (b) The threshold concentrations of the most potent amino acids (arginine, glycine) were between 10(-8) and 10(-9) M. A linear relation between the magnitude of the response and log stimulus concentration held for a wide concentration range for all the amino acids examined. (c) The palatine receptors responded sensitively to various carboxylic acid solutions whose pH was adjusted to neutral. The threshold concentrations varied between 10(-4) and 10(-7) M. The magnitude of the response at 10(-2) M increased with an increase of carbon chain length. (d) The extent of cross-adaptation was examined with various combinations of amino acids. A variety of the response patterns showing complete cross-adaptation, no cross-adaptation, or synergetic interaction was observed. The synergetic interaction was also observed when one amino acid below its threshold concentration was added to the other amino acid below its threshold concentration was added to the other amino acid. No cross-adaptation was observed between amino acids and fatty acids. (e) The treatment of the palate with papain led to loss of the responses to arginine, glycine, and histidine without affecting those to proline and acetic acid. The treatment with pronase E eliminated selectively the response to proline. The possibility that the eel gustatory receptors are responsible for sensing food at a distance was discussed.

Functional expression of pharaonis phoborhodopsin in Escherichia coli.

Pharaonis phoborhodopsin, the photoreceptor of the negative phototaxis of archaebacterial Natronobacterium pharaonis, was functionally expressed in the heterologous system of Escherichia coli. Flash-photolysis on a millisecond time scale indicated that the photochemical properties of ppR expressed in E. coli were the same as those of the native ppR in N. pharaonis. We concluded that the integral membrane protein ppR is correctly folded in vivo in the eubacterial E. coli membrane.