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At least 415 records · Page 23Linked to original sources
PubMed1997

Antiresorptive therapy in hyperthyroid patients: longitudinal changes in bone and mineral metabolism.

The effect of antiresorptive therapy with nasal calcitonin (CT) in recently diagnosed hyperthyroid patients on conventional medical therapy as well as the evolution of bone metabolism were assessed. Forty-five patients with recent-onset hyperthyroidism (<12 weeks) were sex and menopause stratified and randomly allocated to treatment with carbimazole (Neotomizol), carbimazole plus low dose CT (Calsynar; 100 IU/day, 2 days/week), or carbimazole plus high dose CT (Calsynar; 100 IU/day, 14 days/month). Bone mineral density was measured by dual x-ray absorptiometry in lumbar spine, femoral neck, and Ward's triangle at 0, 9, and 18 months of treatment. We also determined free T4, free T3, TSH, osteocalcin, total and bone alkaline phosphatases, tartrate-resistant acid phosphatase, type I collagen C telopeptide, and urinary hydroxyproline every 3 months of follow-up. No significant difference was observed among treatments. A euthyroid state was attained at 3 months. Bone mass increased significantly at the 9 month evaluation (P < 0.05), with a 5-10% net gain during follow-up. Nevertheless, final bone mass was 4-8% smaller than expected. Bone formation markers were increased at 0 and 3 months, with reductions at 6-9 months; resorption bone markers showed a significant reduction at the 3 month evaluation. These results indicate that the euthyroid state partially reduces hyperthyroidism-associated osteopenia, with a bone mass recovery period during the 6-9 early months of effective treatment. This recovery phase is characterized by raised bone formation markers and reduced bone resorption markers. The treatment with nasal CT at the doses assayed has no additional effect over that of attainment of the euthyroid state.

Absorptiometry, Photon↗
PubMed2001

[Man-nature-technology. Philosophical considerations of human-animal-relations].

Man's position in nature is equivocal. Man is part of nature and through evolution is linked to other living beings. Although an "Animal among Animals", nature has provided him with capabilities that separate him from all living beings in its realm. These comprise the ability for reflection and subsequent moral behavior, but also the gift (and necessity) for using technology to shape and subjugate his natural environment. Based upon this "Dual Nature of Man" the essay focuses on the web of interactions between Man, Nature and Technology. Modern ethics of animal protection argue in favor of extending the principle of equality ("To Each His Own") to include animals as well. However, the principle of equality is a purely formal, heuristic rule for assigning morally just determinations of value. It requires normative padding as far as its content is concerned. Consequently, the content of the principle of equality ought to be more closely determined in keeping with the ideal of humanity.

Altruism↗
PubMed2000

Eradication of Helicobacter pylori and stability of remissions in low-grade gastric B-cell lymphomas of the mucosa-associated lymphoid tissue: results of an ongoing multicenter trial.

The normal human stomach is devoid of any organized lymphatic tissue. Acquisition of mucosa-associated lymphatoid tissue (MALT) in the stomach is considered to be a direct consequence of chronic infection with Helicobacter pylori. Thus, MALT appears to be part of the host defense against the pathogen H. pylori. Consequently, lymphomas arising from gastric MALT may be seen as an end point of a clonal evolution starting from the infection. Cumulative data from several studies show that eradication of H. pylori induces complete histologic remissions in about 70%-80% of the patients. Here we present data of an extended analysis of an ongoing multicenter trial. Eighty-four patients with low-grade gastric MALT lymphoma in stage EI were treated using a dual regimen to eradicate H. pylori. Complete remission was observed in 68 (81%) patients; a partial remission was found in seven (8%) patients. In contrast, nine (11%) patients revealed "no change" and were referred for alternative treatment strategies. The majority of these cases were found to harbor high-grade lymphomas in deeper mucosal areas. Polymerase chain reaction (PCR) performed on the VDJ rearrangements of the immunoglobulin heavy chain yielded monoclonal bands in 50 of 65 analyzed patients (77%) at diagnosis. Interestingly, in patients analyzed during follow up after achieving complete histologic remission, ongoing PCR monoclonality was found in 19 of 39 eligible patients (49%). Several patients who developed local relapse of the lymphoma were found in the group with ongoing PCR monoclonality. Together with data from the literature, these results suggest that the majority of low-grade gastric MALT lymphomas in stage EI respond to eradication of H. pylori. Longer follow-up investigations are necessary to determine whether remissions really indicate a cure from the disease and to elucidate whether PCR monoclonality after complete histological remission is predictive of increased relapse rate.

Anti-Bacterial Agents↗
PubMed1997

Structure and function of the low Mr phosphotyrosine protein phosphatases.

Phosphotyrosine protein phosphatases (PTPases) catalyse the hydrolysis of phosphotyrosine residues in proteins and are hence implicated in the complex mechanism of the control of cell proliferation and differentiation. The low Mr PTPases are a group of soluble PTPases displaying a reduced molecular mass; in addition, a group of low molecular mass dual specificity (ds)PTPases which hydrolyse phosphotyrosine and phosphoserine/threonine residues in proteins are known. The enzymes belonging to the two groups are unrelated to each other and to other PTPase classes except for the presence of a CXXXXXRS/T sequence motif containing some of the catalytic residues (active site signature) and for the common catalytic mechanism, clearly indicating convergent evolution. The low Mr PTPases have a long evolutionary history since microbial (prokaryotic and eukaryotic) counterparts of both tyrosine-specific and dsPTPases have been described. Despite the relevant number of data reported on the structural and catalytic features of a number of low Mr PTPases, only limited information is presently available on the substrate specificity and the true biological roles of these enzymes, in prokaryotic, yeast and eukaryotic cells.

Amino Acid Sequence↗
PubMedDate not supplied

Early gene activation in chronic leukemic B lymphocytes induced toward a plasma cell phenotype.

Chronic lymphocytic leukemia (CLL) is characterized by the accumulation of lymphocytes that are arrested at an intermediate stage of B lymphocyte development. CLL B lymphocytes transform (mature) to a plasmacytic phenotype with loss of CD19 and CD20 and the appearance of cytoplasmic immunoglobulin when treated in vitro with phorbol esters. We have used array hybridization technology to describe gene expression patterns for untreated and tetradecanoyl phorbol acetate (TPA)-treated CLL B cells at 5, 10, and 20 min following initial TPA exposure. Three genes, early growth response factor 1 (EGR-1), dual specificity phosphatase 2, and CD69 (early T-cell activation antigen), showed a 2.0-fold or greater increase in mRNA transcription at four or more of six time points in two studies. Upregulation of expression of these genes was confirmed by real-time polymerase chain reaction in the TPA-treated cells of four CLL patients. A progressive increase in gene expression was observed during the 20-min time course for all three genes. In addition, protein expression of EGR-1 and CD69 was increased as measured by immunofluorescence cell analysis. Several genes (PKC, n-myc, jun D, and BCL-2) previously reported as overexpressed in CLL lymphocytes were overexpressed in these studies also, but were not altered by TPA treatment. Genes for proteins whose upregulation requires hours of TPA exposure (the 4F2hc component of the L-system amino acid transporter, prohibition, and hsp60) were assessed, and their later expression contrasted with the early expression of EGR-1, dual specificity phosphatase 2, and CD69. EGR-1 encodes a zinc-finger transcription factor that is induced by pokeweed mitogen and TPA and promotes B lymphocyte maturation. The dual specificity phosphatase 2 encodes an enzyme that reverses mitogen activated protein kinase cell activation by dephosphorylation. The CD69 protein is induced by TPA in thymocytes and is a type II transmembrane signaling molecule in hematopoietic cells. These findings suggest that the products of these three genes may be central to early steps in the TPA-induced evolution of CLL B cells to a plasmacytic phenotype.

Antigens, CD↗
PubMedDate not supplied

Correlation of 99mTc-MIBI brain spect (functional index ratios) and survival after treatment failure in malignant glioma patients.

AIM: This preliminary study was conducted to determine the association between 99mTc-SESTAMIBI (MIBI) brain SPECT and survival in malignant glioma patients after treatment failure. MATERIALS AND METHODS: Twenty-five malignant glioma patients with clinical deterioration were studied. Tomoscintigraphy was performed 15 minutes after intravenous injection of 1110 Mbq MIBI. The images were obtained from a dual head gamma camera using fan beam collimators. Transverse, coronal and sagittal views were reconstructed. Maximum uptake of MIBI in the lesion was expressed as a ratio to that in the controlateral hemisphere. RESULTS: A MIBI uptake was found in all 25 patients. This uptake was correlated to tumour recurrence proved by histological fragments and/or the rapid fatal evolution of these patients. From scintigraphic parameters, two groups of patients were noted. Group 1 (20 patients) had a significantly better survival time (8.25 months) than did Group 2 (5 patients) (2.9 months). Tumour volume predicted survival (group 1 vs group 2, p < 0.0006). A significant correlation between the tumour/background ratio and survival time for each group was observed. CONCLUSION: A correlation between functional index ratios and the time of survival of patients was noted. The functional index ratios of brain SPECT are in accordance with the potential aggressiveness of the recurrent malignant glioma.

Adult↗
PubMed2005

Impact of venlafaxine on gene expression profile in lymphocytes of the elderly with major depression--evolution of antidepressants and the role of the "neuro-immune" system.

Antidepressive drugs offer considerable symptomatic relief in mood disorders and, although commonly discovered by screening with single biological targets, most interact with multiple receptors and signaling pathways. Antidepressants require a treatment regimen of several weeks before clinical efficacy is achieved in patient populations. While the biochemical mechanisms underlying the delayed temporal profile remain unclear, molecular adaptations over time are likely involved. The selective serotonin and noradrenaline reuptake inhibitor, venlafaxine, offers a dual antidepressive action. Its pharmacological behavior, however, is unknown at the genetic level, and it is difficult to monitor in human brain samples. Because the hypothalamic-pituitary-adrenal axis is often severely disrupted in mood disorders, lymphocytes may serve as models of neuropsychiatric conditions. As such, we examined the role of venlafaxine on the gene expression profile of human lymphocytes. DNA microarray was used to measure the expression patterns of multiple genes in human lymphocytes from depressed patients treated with this mood stabilizer. In this self-controlled study, RNAs of control and treated samples were purified, converted into cDNA and labeled with either Cy3 or Cy5, mixed and hybridized to DNA microarrays containing human oligonucleotides corresponding to more than 8,000 genes. Genes that were differentially regulated in response to treatment were selected for follow up on the basis on novelty, gene identity, and level of over-expression/repression, and selected transcripts were profiled by real-time PCR (data have been normalized to beta-actin). Using software analysis of the microarray data, a number of transcripts were differentially expressed between control and treated samples, of which only 57 were found to significantly vary with the "P" value of 0.05 or lower as a result of exposure to venlafaxine. Of these, 31 genes were more highly expressed and 26 transcripts were found to be significantly less abundant. Most selected genes were verified with QRT-PCR to alter. As such, independent verification using QRT-PCR demonstrated the reliability of the method. Genes implicated in ionic homeostasis were differentially expressed, as were genes associated with cell survival, neural plasticity, signal transduction, and metabolism. Understanding how gene expression is altered over a clinically relevant time course of administration of venlafaxine may provide insight into the development of antidepressant efficacy as well as the underlying pathology of mood disorders. These changes in lymphocytes are thought to occur in the brain, and a "neuro-immune system" is proposed by this study.

Aged↗
PubMed2002

[The dual chamber cardiac pacing. A multicenter study apropos of 353 pacemakers].

11 cardiac units, among which 5 experienced cardiac pacing are involved in the study. A retrospective analysis of the operated patient is performed. Authors paid particular attention to the following items: indications, etiologies, per operative parameters, complications and programmatic features of the inserted devices. Demographic date: 176 M, 177 F. The mean age of the sample: 58.4 +/- 16.25 years. The post operative complications occurred in 14% of the patients and were favourable evolution, but the presence of an atrial lead generated the half of the reoperations of which incidence was around 4%. (2%) have led to leave the DDD mode. The favorable results, as well at the technical levels as the evaluating level, witness that the tunisian centers have acquired a sufficient knowledge in the field of the conventional pacing and could now born on other problems, out passing the field of the bradycardias.

Adult↗
PubMed2003

Evolution of the immunomodulatory role of the heat shock protein gp96.

In mammals, certain heat shock proteins (hsps) participate in specialized responses to stressors associated with pathogens or tumors, and as such, act as agents of immune surveillance interacting with both innate and adaptive immunity. We are investigating the conservation of this role throughout the class of vertebrates. We have shown that in Xenopus as in mammals, gp96, the major resident of the endoplasmic reticulum, generates MHC-restricted thymus-dependent immunity in vivo and CR in vitro against minor histocompatibility (H) antigens. By as yet unclear mechanisms that may involve classical MHC-unrestricted cytotoxic CD8+ T cells, gp96 also elicits peptide-specific responses against MHC-class I-negative tumors in adult frogs that may involve cytotoxic NK, MHC-unrestricted CD8+ T and NK/T cells. In naturally MHC class I-deficient but immunocompetent Xenopus larvae, gp96 also generates an innate type of anti-tumor response that is independent of chaperoned peptides. Finally, in a subset of Xenopus sIgM+ B cells, a substantial fraction of gp96 is directed to the cell surface by an active process that is upregulated by bacterial stimulation. This may allow gp96 to access the extracellular compartment without necrosis. Given the dual abilities of gp96 to chaperone antigenic peptides and to modulate innate immune responses, we propose that stimulated B cells that are up-regulating surface gp96 can directly interact with antigen presenting cells (APC) and/or T helper (Th) cells to trigger or amplify immune responses.

Adjuvants, Immunologic↗
PubMed1999

Melatonin modulation of prolactin and gonadotrophin secretion. Systems ancient and modern.

Recent studies in sheep indicate that the pineal melatonin signal which transduces effects of photoperiod acts at separate sites in the pituitary gland and brain to regulate seasonality in prolactin (PRL) and gonadotrophin secretion. The pituitary gland is the proposed site for control of PRL based on the observation that hypothalamo-pituitary disconnected (HPD) rams continue to show normal patterns of PRL secretion in response to changes in photoperiod or treatment with melatonin. Lactotrophs do not express melatonin receptors, thus this pituitary effect is assumed to be mediated by cells in the pars tuberalis via "tuberalin". The mediobasal hypothalamus (MBH) is the putative target for gonadotrophin control since: i) gonadotrophin secretion is dependent on pulsatile GnRH secretion from the MBH, ii) local administration of melatonin in the MBH, but not in other areas of the brain and pituitary gland, readily reactivates GnRH-induced LH and FSH secretion in photo-inhibited rams; and iii) treatment of HPD rams with a chronic pulsatile infusion of GnRH stimulates gonadotrophin secretion irrespective of photoperiod. Complementary studies conducted by others in the Syrian hamster, have shown that lesions in the MBH block the action of melatonin on gonadotrophin but not on prolactin secretion; this supports the "dual-site hypothesis". Since all photoperiodic mammals are essentially similar in hyper-secreting PRL under long days, the pituitary control mechanism for PRL is regarded as conserved (ancient) with the pleiotrophic actions of PRL inducing a summer physiology (e.g. growth of summer pelage). In contrast, the variation between species in the timing of the gonadal cycle indicates that evolution has independently modified the melatonin-sensitive neural circuits in the MBH to permit the species-specific timing of the mating season.

Animals↗
PubMed2005

A dynamical analysis of tennis: concepts and data.

This research reported here draws on self-organization theories and dynamical system models to investigate the collective behaviour of tennis players. In tennis, the unceasing to-and-fro displacements of a player about a "home" reference position, located in the middle of the baseline, are akin to those of an oscillator, and the reciprocal attending of both players establishes an informational linkage. Thus, theoretically, the displacement of the two players can be analysed as a system formed by two coupled non-linear oscillators. In such a system, relative phase has been shown to be an adept measure of the temporal synchronization between the oscillators. We hypothesized that relative phase is a relevant collective variable to characterize the relative motion of tennis players. Four players were videotaped and their displacements analysed. The results revealed just two stable patterns of synchronization, in-phase and anti-phase, as the players moved in the same or opposite directions, respectively. Moreover, relative phase showed two types of evolution within trials: either it remained stable at in-phase or anti-phase, or it exhibited transitions between these two modes. In accordance with our hypothesis, the results identified relative phase as a pertinent collective variable to represent both invariance and change in the relative displacements of tennis players. Such a finding opens new avenues for investigating dual sports.

Biomechanical Phenomena↗
PubMed2003

Bioavailable estradiol and an aromatase gene polymorphism are determinants of bone mineral density changes in men over 70 years of age.

The question of whether and to what extent the sex steroid deficiency in elderly men contributes to the pathogenesis of bone loss has not been fully explored. The aim of the present study was to assess the association of serum bioavailable (Bio) estradiol (E(2)) with the evolution of bone mineral density (BMD) in 214 community-dwelling men aged 71-86 yr as well as the possible modulation of estrogen effects by a tetranucleotide (TTTA)(n)-repeat polymorphism of the CYP19 gene, which encodes the aromatase enzyme that converts androgens into estrogens. BMD was measured at yearly intervals over a period of 4 yr using dual x-ray absorptiometry. Fasting blood was analyzed at baseline for testosterone (T), E(2), and SHBG; the respective bioavailable fractions, BioT and BioE(2), were calculated. Serum BioE(2) was associated with baseline BMD at different assessed skeletal sites, with correlation coefficients ranging between 0.23 and 0.37 (P < 0.001). Estimated annual percentage change of BMD (%BMD) was -0.39% [95% confidence index (CI), -0.56, -0.22] at the total hip, -0.04% (95% CI, -0.29, 0.21) at the femoral neck, and -0.37% (95% CI, -0.45, -0.29) at the total distal forearm. Higher circulating BioE(2) levels were associated with less bone loss at the forearm and the hip (P < 0.05). The CYP19 gene (TTTA)(n)-repeat length (determined by fragment analysis) was not associated with baseline BMD in the total group of elderly men. However, a significant association was observed between the CYP19 genotype and BMD change at the distal forearm; the highest bone loss was observed in subjects homozygotic for the shortest observed allele length of (TTTA)(7)-repeats (P < 0.02). The CYP19 (TTTA)(n)-repeat length was not associated with either baseline BioE(2) or the BioT/BioE(2) ratio. In multiple linear regression models, the CYP19 genotype and serum BioE(2) were determinants of %BMD change at the forearm (P < 0.05). No significant contribution of BioT to %BMD change was evident. As to fracture risk, the allele containing the shortest (TTTA)(n)-repeat length was more represented not only in elderly men with a positive personal fracture history (Pearson's chi(2) test = 4.03; df = 1; P = 0.05) but also in study subjects with a positive fracture history in their first-degree relatives (Pearson's chi(2) test = 6.48; df = 1; P = 0.01). In conclusion, the results of this prospective observational study support the view that BioE(2) is a determinant of bone density changes in elderly men and, furthermore, provide an indication that the aromatase enzyme may exert a direct modulatory action on bone metabolism at the tissue level in elderly men.

Aged↗
PubMed2000

The crystal structure of a sulfurtransferase from Azotobacter vinelandii highlights the evolutionary relationship between the rhodanese and phosphatase enzyme families.

Rhodanese is an ubiquitous enzyme that in vitro catalyses the transfer of a sulfur atom from suitable donors to nucleophilic acceptors by way of a double displacement mechanism. During the catalytic process the enzyme cycles between a sulfur-free and a persulfide-containing form, via formation of a persulfide linkage to a catalytic Cys residue. In the nitrogen-fixing bacteria Azotobacter vinelandii the rhdA gene has been identified and the encoded protein functionally characterized as a rhodanese. The crystal structure of the A. vinelandii rhodanese has been determined and refined at 1.8 A resolution in the sulfur-free and persulfide-containing forms. Conservation of the overall three-dimensional fold of bovine rhodanese is observed, with substantial modifications of the protein structure in the proximity of the catalytic residue Cys230. Remarkably, the native enzyme is found as the Cys230-persulfide form; in the sulfur-free state the catalytic Cys residue adopts two alternate conformations, reflected by perturbation of the neighboring active-site residues, which is associated with a partly reversible loss of thiosulfate:cyanide sulfurtransferase activity. The catalytic mechanism of A. vinelandii rhodanese relies primarily on the main-chain conformation of the 230 to 235 active-site loop and on a surrounding strong positive electrostatic field. Substrate recognition is based on residues which are entirely different in the prokaryotic and eukaryotic enzymes. The active-site loop of A. vinelandii rhodanese displays striking structural similarity to the active-site loop of the similarly folded catalytic domain of dual specific phosphatase Cdc25, suggesting a common evolutionary origin of the two enzyme families.

Amino Acid Sequence↗
PubMed1997

What role does Helicobacter pylori eradication play in gastric MALT and gastric MALT lymphoma?

The concept of mucosa-associated lymphoid tissue (MALT) has been introduced to differentiate biological functions from behavior of nonnodal vs. nodal lymphoid tissues. Lymphomas arising from MALT also behave differently than typical nodal lymphomas. In contrast to other tissues, MALT in the stomach is almost exclusively a result of Helicobacter pylori infection. Thus, MALT is part of the host defense against the pathogen H. pylori. Consequently, lymphomas arising from gastric MALT may be a clonal evolution starting from the infection. In low-grade gastric MALT lymphoma, cure of the infection may induce complete histological remission in the majority of patients. Investigators have recently reported that complete remission rate is between 70% and 80%. In an extended analysis, we have treated 84 patients with low-grade gastric MALT lymphoma in stage El, using a dual regimen to eradicate H. pylori. Complete remission was observed in 68 (80%) patients; a partial remission was found in 4 patients. In contrast, 12 patients showed no change and were referred to alternative treatment. In patients in complete remission, a polymerase chain reaction assay for the rearranged immunoglobulin heavy-chain gene remained positive in many cases. Together with data from the literature, these data suggest that the majority of patients with low-grade gastric MALT lymphomas in stage El respond to eradication of H. pylori. Longer follow-up investigations are necessary to determine if remissions indicate a cure from the disease.

Gastric Mucosa↗
PubMed1995

Monitoring electrocardiographic changes: results of a national survey.

This article describes the electrocardiographic (ECG) monitoring practices of a national sample of nurses caring for patients who have suffered an acute myocardial infarction (AMI). Of specific interest were lead section practices, diagnosis of infarct evolution, and use of right precordial leads. Two populations were randomly sampled in an attempt to minimize sampling bias. Of the 2,549 mailed surveys, 882 were returned (34.8% response rate) from 48 states. Lead II was used most commonly (66.3%) with a single-channel system. Lead II plus V1 (or MCL1) was selected most often (62.4%) with dual-channel systems. Few (20%) respondents reported "always" modifying lead selection based on location of AMI; selected most often (62.4%) with only 43.3% correctly identified classic ischemia, injury, and infarction. Right precordial leads are used infrequently in practice.

Chi-Square Distribution↗
PubMed2006

Numerical analysis and experimental results of high-power Er/Pr:ZBLAN 2.7 microm fiber lasers with different pumping designs.

Gain factor and output performance of erbium-praseodymium codoped ZBLAN double-clad fiber lasers at 2.7 microm with different pumping designs were calculated and analyzed. Single-end backward pumping with a highly reflective mirror butted against one fiber end and dual-end pumping with Fresnel reflections from both fiber ends were found to be the most efficient pumping designs. Ten-watt-level Er/Pr:ZBLAN fiber lasers proved to be achievable with recent diode laser and ZBLAN fiber technologies. Their corresponding optimum fiber lengths for different pumping configurations were determined. It was also found that fiber lasers with a flat evolution of gain factor can obtain the largest output power. Experimental results of 4 m and 12 m fiber lasers showed very good agreement with simulation results.

Journal Article↗
PubMed1979

Evolutionary relationships of the gut hormones.

Peptides identical or related to mammalian gut hormones occur widely, not just in gut endocrine cells but also in central or peripheral nerves, amphibian skin glands, and a variety of invertebrate tissues. The dual distribution in brain and gut was probably already established early in the vertebrate line; representatives of the oldest vertebrate group, the cyclostomes, have cholecystokinin-like factors in gut endocrine cells and in brain. The related sequences of certain gut peptides, notably gastrin and cholecystokinin (CCK), and secretin, glucagon, vasoactive intestinal polypeptide (VIP), and gastric inhibitory peptide (GIP), indicate evolution from common ancestral molecules by gene duplication and divergence. Functionally important residues are conserved. Thus the COOH-terminal pentapeptide common to gastrin and CCK also contains their minimal active fragment. There are also evolutionary changes at the level of the target organ receptor mechanisms: these are also evolutionary changes at the level of the target organ receptor mechanisms; these are illustrated by evidence suggesting that secretin regulates the flow of pancreatic juice in mammals whereas the structurally related peptide VIP has a similar role in birds.

Animals↗
PubMed1978

Resonance Raman spectroscopy of the retinylidene chromophore in bacteriorhodopsin (bR570), bR560, M421, and other intermediates: structural conclusions based on kinetics, analogues, models, and isotopically labeled membranes.

Resonance Raman spectra of various intermediates in the bacteriorhodopsin proton pumping cycle have been obtained at physiological and low temperatures. To interpret these data, spectra of modél compounds, bacteriorhodopsin analogues, and isotopically labeled membranes have been measured. These results demonstrate that a protein group interacts with the Schiff base proton and, thus, the chromophore in protonated bacteriorhodopsin species is not a simple protonated Schiff base. This accounts for the abnormally low frequency of the C=N+H vibrational mode in bacteriorhodopsin and other failures to model the chromophore in bR570 with a simple butylamine protonated Schiff base of all-trans-retinal. To obtain the resonance Raman spectrum of M412 at physiological pH and temperatures, a dual beam kinetic technique was developed. We demonstrate that in the fingerprint region of the resonance Raman spectrum M412 is modeled accurately by a simple unprotonated butylamine Schiff base of all-trans-retinal. Spectral resolution and the solution environment of the membrane suspensions play important roles in this conclusion. Kinetic resonance Raman techniques are also used to monitor the time evolution of the M412 species and the intermediates which precede it. We find spectral features in our kinetic data which can be assigned to L550, and we present evidence for a new unprotonated species (X) which occurs before M412. Single pass flow resonance Raman spectra of bR560 also have been obtained, and, although bR570 and M412 appear to have all-trans chromophores, there are 13-cis-like features in the spectra of bR560, L550, and X.

Bacteriorhodopsins↗