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M Ferrand

Publications and source records attributed to M Ferrand.

At least 19 recordsLinked to original sources
PubMed1995

Structural model of the photosynthetic reaction center of Rhodobacter capsulatus.

The reaction center (RC) from the photosynthetic bacterium Rhodobacter (Rb.) capsulatus has been the subject of a considerable amount of molecular biological and spectroscopic work aimed at improving our understanding of the primary steps of photosynthesis. However, no three-dimensional structure is available for this protein. We present here a model obtained by combining information from the structure of the highly homologous RC from Rhodopseudomonas (Rps.) viridis with molecular mechanics and simulated annealing calculations. In the Rb. capsulatus model the orientations of the bacteriochlorophyll monomer and the bacteriopheophytin on the branch inactive in electron transfer differ significantly from those in the RCs of Rps. viridis and Rb. sphaeroides. The bacteriopheophytin orientational difference is in good accord with previous linear dichroism measurements. A comparison is made of interactions between the pigments and the protein environment that may be of functional significance in Rps. viridis, Rb. sphaeroides, and Rb. capsulatus.

Amino Acids↗
PubMed1994

Bispecific thyroglobulin and thyroperoxidase autoantibodies in patients with various thyroid and autoimmune diseases.

Recent evidence indicates that thyroid autoimmune disorders are associated with the presence of circulating autoantibodies (aAb) with dual specificity for thyroglobulin (TG) and thyroperoxidase (TPO). The question of whether these aAb, called TGPO aAb, are of clinical relevance compared to TG and TPO aAb remains to be determined. The availability of purified preparations of human TG and TPO allowed the development of a specific and sensitive RIA for TGPO aAb in serum. In the present study, we compared levels of aAb that cross-react with both TG and TPO (TGPO aAb) and total TG and TPO aAb levels, respectively, in sera from 84 normal controls and 226 patients with various thyroid and autoimmune diseases, including nontoxic goiter (n = 50), toxic nodular goiter (n = 13), thyroid carcinoma (n = 20), primary idiopathic myxedema (n = 15), postpartum thyroiditis (n = 11), Hashimoto's thyroiditis (n = 38), pernicious anemia (n = 27), rheumatoid arthritis (n = 19), and insulin-dependent diabetes mellitus (n = 33). In addition, 16 patients with Hashimoto's thyroiditis were studied before therapy and after more than 3 months of treatment with L-T4. It was shown that TGPO aAb were generally, but not always, present in the serum of patients with Hashimoto's thyroiditis, which also contained TG and TPO aAb. In contrast, TGPO aAb were undetectable in normal controls (excepting a few cases reaching borderline levels) as well as in sera from the majority of the other patients tested. Selecting sera positive for TGPO and either TG or TPO aAb, a statistically significant correlation was found between TGPO and TG (n = 26; P < 0.005), but not TPO aAb. Interestingly, the TGPO aAb level significantly decreased in patients with Hashimoto's thyroiditis after hormonal therapy (P < 0.05), some of them shifting from TGPO aAb positive before treatment to negative after treatment. In conclusion, TGPO aAb determination distinguishes Hashimoto's patients from patients with either thyroid and/or autoimmune diseases. The specific presence of TGPO aAb in a subset of Hashimoto's patients and their variation during T4 therapy remain to be understood. This could give a clue to mechanisms of autoimmune thyroid disease.

Adolescent↗
PubMed1994

Autoantibodies and monoclonal antibodies directed to an immunodominant antigenic region of thyroglobulin interact with thyroperoxidase through an interspecies idiotype.

We investigated whether thyroglobulin (TG) autoantibodies (aAb) cross-react with thyroperoxidase (TPO) through an idiotypic structure using pooled normal human IgG (NhlgG) as a natural anti-idiotype reagent. Affinity-purified TG aAb from pooled IgG of patients with autoimmune thyroid disease were chromatographed on Sepharose-bound NhlgG. About one fourth of the loaded material bound to and eluted from the coupled gel. Eluted TG aAb were found reactive to TG and TPO and their TPO but not TG binding was strongly inhibited by molar excess of NhlgG. These TG aAb appeared to be mainly directed to an immunodominant TG antigenic region defined by TG monoclonal antibodies (mAb) from a single cluster of reactivity. These TG mAb were also found to recognize TPO and their binding to TPO but not TG was inhibited by molar excess of NhlgG as already observed with TG aAb. Taken together, these results indicated that TPO interacts with an idiotype present on human TG aAb and mouse TG mAb displaying a similar epitopic specificity; this interspecies idiotype is recognized by anti-idiotype antibodies present in NhlgG. Our results suggest that thyroid autoimmunity can be envisaged, at least in part, as a disturbance in interconnected idiotypic networks.

Adult↗
PubMed1993

Thermal motions and function of bacteriorhodopsin in purple membranes: effects of temperature and hydration studied by neutron scattering.

The internal dynamics of bacteriorhodopsin, the light-driven proton pump in the purple membrane of Halobacterium halobium, has been studied by inelastic neutron scattering for various conditions of temperature and hydration. Light activation can take place when the membrane is vibrating harmonically. The ability of the protein to functionally relax and complete the photocycle initiated by the absorption of a photon, however, is strongly correlated with the onset of low-frequency, large-amplitude anharmonic atomic motions in the membrane. For a normally hydrated sample, this occurs at about 230 K, where a dynamical transition from a low-temperature harmonic regime is observed. In moderately dry samples, on the other hand, in which the photocycle is slowed down by several orders of magnitude, no transition is observed and protein motions remain approximately harmonic up to room temperature. These results support the hypothesis, made from previous neutron diffraction studies, that the "softness" of the membrane modulates the function of bacteriorhodopsin by allowing or not allowing large-amplitude motions in the protein.

Bacteriorhodopsins↗
PubMed1993

Structure and dynamics of bacteriorhodopsin. Comparison of simulation and experiment.

Global features of the structure and dynamics of bacteriorhodopsin are investigated using molecular modelling, dynamical simulations and neutron scattering experiments. The simulations are performed on a model system consisting of one protein molecule plus intrinsic water molecules. The simulation-derived structure is compared with neutron diffraction data on the location of water and with the available electron microscopy structure of highest resolution. The simulated water geometry is in good accord with the neutron data. The protein structure deviates slightly but significantly from the experiment. The low-frequency vibrational frequency distribution of a low-hydration purple membrane is derived from inelastic neutron scattering data and compared with the corresponding simulation-derived quantity.

Bacteriorhodopsins↗
PubMed1993

Significance of thyroglobulin antibodies cross-reactive with thyroperoxidase (TGPO antibodies) in individual patients and immunized mice.

Thyroglobulin (TG) and thyroperoxidase (TPO), both involved in thyroid hormone synthesis, represent major autoantigens in thyroid autoimmune disease. Despite numerous studies, the emergence, pathophysiological significance and role of autoantibodies to TG and TPO remain elusive. The recent identification of a new category of thyroid-specific autoantibody interacting with both TG and TPO (TGPO autoantibodies) offers a new opportunity in the study of thyroid autoimmunity. To gain a better insight into the significance of these TGPO autoantibodies, measurement in individual samples appeared necessary. The unique property of TGPO autoantibodies, simultaneous binding to TG and TPO, was used to set up a sandwich method which combined coated TG and radio-iodinated TPO. This method was found to be strictly specific for TGPO autoantibodies and sensitive enough to assay TGPO autoantibodies in serum. In humans, TGPO autoantibodies were found in most of the sera with high TG and TPO autoantibody titres, but not in sera negative for TG autoantibodies, whatever the TPO autoantibody titre. Furthermore, high TGPO autoantibody titres were found in sera strongly cytotoxic for cultured porcine thyroid cells. However, significant correlation of TGPO autoantibody titre was observed neither with TG and TPO autoantibody titres (n = 48) nor with complement-dependent cytotoxicity (n = 50). TGPO antibody assay was also performed in individual plasma of CBA/J mice immunized with either human TG (n = 6) or human TPO (n = 6). Immunization with TG induced high levels of not only TG but also TGPO antibodies, which exhibited a strong reactivity for TPO and whose binding to TG and TPO was fully inhibited by TG. In contrast, immunization with TPO induced high levels of only specific TPO antibodies accompanied by low levels of specific TG antibodies. In this case TGPO antibodies were not detected. Of note, TG- and TPO-immunized mice mounted an immune response against their own TG, but did not exhibit histological signs of thyroiditis. Large panels of TG and TPO MoAbs were also investigated with this method: 18/25 TG MoAbs and only 1/13 TPO MoAbs were found cross-reactive. Taken together, these data provide evidence that TGPO antibodies are effectively present in individual patients and TG-immunized mice, are different from specific TG and TPO antibodies, and may derive from natural B cell repertoire by autoimmune processes involving TG and not TPO.

Animals↗
PubMed1992

Immunopurification and characterization of thyroid autoantibodies with dual specificity for thyroglobulin and thyroperoxidase.

The presence of autoantibodies (aAbs) to thyroglobulin (TG) and thyroperoxidase (TPO) in most of the patients with autoimmune thyroid disease is now well documented. Studies of these aAbs suggested that some, termed TGPO aAbs, could interact with both TG and TPO. This hypothesis was investigated using IgG fraction from a pool of 25 patients' sera with high TG and TPO aAb titres. Immunopurification of TG, TPO and TGPO aAbs was carried out by sequential affinity chromatography using a large quantity of highly purified human TG and TPO. TGPO aAbs, obtained absorption-elution of affinity purified TG aAbs onto a TPO column, were found to represent about 20% of the TG reactive aAbs and 0.23% of the total amount of IgG. Purified TGPO aAbs were characterized and compared to specific TG and TPO aAbs. In contrast to TG and TPO aAbs which recognized only their target antigen, TGPO aAbs showed high affinity interactions with both TG and TPO. As compared to TG aAbs, TGPO aAbs displayed similar affinity for native TG and higher affinity for denatured TG. Compared to TPO aAbs, TGPO aAbs showed lower affinity for both native and denatured TPO. TGPO aAbs also differed from specific TG and TPO aAbs with regard to IgG subclass distribution and antigen fine specificities as determined by monoclonal antibody assisted mapping of TG and TPO surface epitopes. Taken together, these data indicate that TGPO aAbs are effectively present in the serum of patients with autoimmune thyroid disease. TGPO aAbs may be considered as a subpopulation of TG aAbs with the unique property to cross-react with TPO. The existence of aAbs cross-reacting with these functionally and antigenically related thyroid molecules could lead to a re-examination of the emergence of thyroid autoimmunity.

Antibody Specificity↗
PubMed1992

[Ultrastructural changes in nucleoli during embryogenesis of Marsilea vestita].

The nucleoli of oospheres and young embryos of Marsilea vestita exhibited a typical ultrastructure with segregation into several compact nucleolus-like bodies and small granules of similar appearance. When embryos reached the 16 cell stage, the granules disappeared and the nucleolar-like bodies changed into classical nucleolar components. When the embryos were submitted to hypothermic shock, the nucleoli returned to the characteristic ultrastructure of the very young stages. In ageing oospheres the nucleus was full of very compact clumps. No detectable RNA synthesis occurred in the oospheres and young embryos, and the same phenomenon applied in ageing oospheres and after cold shock treatment. Transcriptional activity started with the 8-16 cell stage in the reference embryos. These results suggest that nucleolar ultrastructure is correlated with rRNA synthesis.

Cell Nucleolus↗
PubMed1991

Stored mRNA in early embryos of a fern Marsilea vestita: a paternal and maternal origin.

In early embryos of the fern Marsilea vestita, we have shown that transcription did not begin until the 8-cell stage, even though protein synthesis was taking place. As in animal species, the pre-existence of stored maternal mRNA has been suspected in embryonic cells. To investigate the presence of such long-lived mRNA we have used in situ hybridization method using [3H]polyuridylic acid, [3H]poly(U), as a probe. Temporal and spatial changes in the distribution of poly(A)+ RNA were followed throughout all the different phases of embryogenesis. Poly(A)+ RNA was present in the cytoplasm of both unfertilized and fertilized eggs and early embryos, while the nucleus exhibited no, or a moderate, [3H]-poly(U) binding activity. New poly(A)+ RNA was detected in the nucleus only at the 16-cell stage, when cytological differentiation of the root apical cell became morphologically detectable. After in situ hydridization with the labeled probes, the numerous silver grains detected autoradiographically over the mature sperm, indicate the presence of poly(A)+ RNA molecules associated with the male genome. We discuss the possible role of poly(A)+ RNA of maternal origin in supporting early translation prior to synthesis of new mRNA. Additional studies are needed to elucidate the role of stored paternal mRNA.

Autoradiography↗
PubMed1989

Relationship between immunological structure and biochemical properties of human thyroid peroxidase.

Although the primary structure of human thyroid peroxidase (hTPO) has been recently deciphered, little is known about its spatial conformation. Such information is of crucial importance in any attempt to relate the structure with the function of hTPO. To probe the antigenic surface of hTPO and to correlate its immunological structure to its biochemical properties, we used 13 monoclonal antibodies (mAb) displaying various affinity for hTPO. Criss-cross experiments showed 7 clusters of reactivity which were interpreted as reflecting 7 epitopes on the surface of the hTPO molecule. Extending our analysis to partial and nonsymmetrical cross-reactivities, these epitopes were shown to be localized in 4 antigenic domains of the hTPO. We further investigated the nature of these 7 hTPO epitopes by testing mAb binding to peroxidases from various origins and chemically modified hTPO; 3 epitopes were shown to be evolutionary conserved, and 5 resistant to reduction and denaturation. We also analyzed the role of the hTPO epitopes in the enzymatic activity and autoimmune targeting of the molecule. Nine epitopes were shown to be localized at the vicinity of both catalytic sites as the binding of their respective mAb modulated the enzyme activity. Autoantibodies from patients presenting with autoimmune thyroid disorders were essentially directed to epitopes similar or adjacent to those recognized by 8 of the 13 mAb and present on only 2 antigenic domains of hTPO. Taken together these data allowed us to propose a tentative map of the surface of the hTPO molecule which associates its epitopic structure with its biochemical functions.

Antibodies, Monoclonal↗
PubMed1988

Novel routine assay of thyroperoxidase autoantibodies.

This radioimmunoassay was developed for specific and large-scale routine measurement of autoantibodies to thyroperoxidase (TPO), an enzyme recently identified as the thyroid microsomal antigen. Because of the scarcity of purified thyroperoxidase, we did not base the assay on the antigen-coated method but rather on autoantibody inhibition of the binding of labeled TPO to a solid-phase-bound monoclonal antibody to TPO. This assay design ensured highly specific measurements without interference from irrelevant thyroid antigens and autoantibodies. When we used affinity-purified autoantibodies to TPO as standards, the range of the curve extended over 10(3)-fold differences in the autoantibodies' concentrations, which allowed us to assay most sera without dilution. Within- and between-assay coefficients of variation (CVs) ranged from 6.1% to 11.5% and from 6.6% to 12.0%, respectively. The correlation between anti-TPO and antimicrosomal autoantibodies, as assessed by hemagglutination test, was highly significant (r = 0.90, P less than 0.0001). This assay is sensitive, easy to perform, and requires only trace amounts of purified TPO.

Antibodies, Monoclonal↗
PubMed1987

Thyroid peroxidase is the organ-specific 'microsomal' autoantigen involved in thyroid autoimmunity.

Autoantibodies (aAb) in serum of patients with autoimmune thyroid diseases (AITD) are directed to an antigen associated with thyroid microsomes. Although it has been investigated over almost three decades, the nature of this autoantigen remained unknown. Taking advantage of monoclonal antibodies (mAb) produced in our laboratory, we have demonstrated that thyroid peroxidase (TPO) is the 'microsomal' antigen. Sera of patients with AITD strongly inhibited the binding of only one of 19 mAb raised against human thyroid plasma membranes. This mAb did not react with thyroglobulin but achieved significant binding to preparations of human, bovine and porcine TPO, bovine lactoperoxidase and human myeloperoxidase without altering the enzyme activity. The mAb has been used to immunopurify the human TPO from solubilized thyroid microsomes. The procedure allowed high purification (approximately 3500-fold) of the native enzyme with a reasonable yield (approximately 10 mg TPO/kg thyroid tissue). Human TPO exhibited a specific activity of 350-400 guaiacol U/mg, a peak in the Soret region and a ratio of A411 nm to A280 nm of 0.20-0.25. Upon SDS-polyacrylamide gel electrophoresis, the purified enzyme gave two contiguous bands in the 100 kDa region. Performed in non-reducing conditions, electrophoresis of TPO showed one band in the same 100 kDa region. Sera with aAb to the microsomal antigen immunoprecipitated purified TPO to an extent ranging from 80 to 100% of the initial enzyme amount while sera from normal subjects or from patients with undectable level of anti-microsomal aAb elicit a decrease of less than 30% of the total TPO activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed1986

Interaction of highly purified thyroid peroxidase with anti-microsomal antibodies in autoimmune thyroid diseases.

Among the several antigens involved in autoimmune thyroid diseases, the microsomal antigen has been recently identified as the human thyroid peroxidase (TPO). The availability of highly purified TPO and anti-TPO monoclonal antibody (mAb) allowed us to study in more details anti-TPO autoimmune antibodies (aAb) and their relationship with anti-microsomal aAb. Only sera with anti-microsomal aAb, as assayed by passive hemagglutination, highly immunoprecipitated purified TPO; anti-thyroglobulin aAb did not contribute to this effect. IgG binding to TPO and inhibition of mAb binding to TPO were also observed using anti-microsomal positive and anti-thyroglobulin negative sera. The correlation between anti-TPO and anti-microsomal aAb titers was found significant (p less than 0.02). These data suggest that anti-microsomal and anti-TPO immunoreactivity are presented by the same aAb. It could not be ruled out, however, that some of the anti-microsomal aAb detected by passive hemagglutination could be directed against antigens different from TPO. The use of highly purified TPO and anti-TPO mAb would allow large scale studies of anti-TPO aAb, necessary for investigating their pathological significance in thyroid disorders.

Antibodies, Monoclonal↗
PubMed1985

Purification of the human thyroid peroxidase and its identification as the microsomal antigen involved in autoimmune thyroid diseases.

Human thyroid peroxidase (TPO) has been purified from thyroid microsomes by immunoaffinity chromatography using a monoclonal antibody (mAb) to TPO. The eluted material had a specific activity of 381 U/mg and exhibited a peak in the Soret region. The ratio of A411 to A280 ranged from 0.20 to 0.25. Upon SDS-polyacrylamide gel electrophoresis, the purified enzyme gave two contiguous bands in the 100 kDa region. Further, it has been demonstrated that sera with anti-microsomal autoantibodies from patients presenting Graves' or Hashimoto's thyroiditis diseases were able to bind to purified TPO and to inhibit in a dose-dependent manner the mAb binding to purified TPO. This suggests that TPO is the thyroid antigen termed to date the microsomal antigen.

Antibodies, Monoclonal↗
PubMed1985

Cell membrane coating with glutaraldehyde: application to a versatile solid-phase assay for thyroid membrane proteins and molecules interacting with thyroid membranes.

In defined conditions, glutaraldehyde was shown to tightly bind cell membranes to flexible microtiter plates without significant alteration of the antigenic and functional properties of membrane proteins. In the presence of 0.06% glutaraldehyde, human thyroid membranes were bound to plastic firmly enough to resist numerous washing and flicking steps; the coated membranes remained almost unaltered with regard to monoclonal antibody and thyrotropin binding as well as adenylate cyclase and peroxidase activities. Based on the use of thyroid membrane-coated microtiter plates, a versatile solid-phase assay was developed which allowed screening of anti-membrane monoclonal antibodies, detection of thyrotropin-displacing activity in hormone and antibody preparations, and monitoring of fractionation experiments of solubilized membrane antigens and thyrotropin receptor. It was concluded that the use of glutaraldehyde for coating cell membranes to flexible microtiter plates enabled the establishment of simple, rapid, and reliable assays for detection and quantitation of membrane proteins and molecules interacting with membranes.

Adenylyl Cyclases↗
PubMed1985

[Antigenic relation between thyroid peroxidase and the microsomal antigen implicated in auto-immune diseases of the thyroid].

Pools of sera from patients with Graves' disease or Hashimoto's thyroiditis highly inhibit the binding to human thyroid membranes of one of 19 monoclonal antibodies raised against preparations of human thyroid membranes. This monoclonal antibody reacts with human and bovine thyroid peroxidase and bovine lactoperoxidase but not with human hemoglobin, cytochrome c and other related molecules. These results indicate that the thyroid peroxidase and the microsomal antigen are antigenically related. These data taken together with those from other groups, highly suggest that thyroid peroxidase is the microsomal antigen involved in autoimmune thyroid diseases.

Adult↗