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M Genest

Publications and source records attributed to M Genest.

At least 19 recordsLinked to original sources
PubMed2006

Lateral mobility of proteins in liquid membranes revisited.

The biological function of transmembrane proteins is closely related to their insertion, which has most often been studied through their lateral mobility. For >30 years, it has been thought that hardly any information on the size of the diffusing object can be extracted from such experiments. Indeed, the hydrodynamic model developed by Saffman and Delbrück predicts a weak, logarithmic dependence of the diffusion coefficient D with the radius R of the protein. Despite widespread use, its validity has never been thoroughly investigated. To check this model, we measured the diffusion coefficients of various peptides and transmembrane proteins, incorporated into giant unilamellar vesicles of 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC) or in model bilayers of tunable thickness. We show in this work that, for several integral proteins spanning a large range of sizes, the diffusion coefficient is strongly linked to the protein dimensions. A heuristic model results in a Stokes-like expression for D, (D proportional, variant 1/R), which fits literature data as well as ours. Diffusion measurement is then a fast and fruitful method; it allows determining the oligomerization degree of proteins or studying lipid-protein and protein-protein interactions within bilayers.

Bacterial Outer Membrane Proteins↗
PubMed2002

[Differential therapy in radial head fractures].

The therapeutic regimen of radial head fractures, especially of displaced and comminuted types is controversial. The radial head resection has been critically reviewed over the past years. From 1984-1993 and 1996-1999, 105 radial head fractures were treated in our hospital. 74 were subject to clinical and radiological follow-up. Fracture-types were classified according to Mason. Undisplaced fractures were treated conservatively, displaced 2-fragment-fractures by an open reduction and screw fixation, and multifragment-fractures by a radial head resection. The results were studied on a functional and radiological basis using the "Functional Rating Index" of Broberg and Morrey and the radiological Score of Albrecht and Ganz. After conservative therapy over 80 % achieved excellent and good as well as 12.5 % satisfactory and 6.3 % unsatisfactory results. After reduction and internal fixation again 80 % had excellent and good results. After radial head resection excellent and good results were achieved in 54.6 % of the cases, satisfactory results in 24.2 % and in 21.2 % unsatisfactory results, however prognosis-influencing concomitant injuries were often present in the latter group. Using the right indication and technique, the radial head resection still is a recommendable therapeutic procedure with an altogether good prognosis. This especially applies to isolated radial head fractures where excellent and good results can be achieved in approximately 70 %.

Adolescent↗
PubMed2001

Self diffusion and spectral modifications of a membrane protein, the Rubrivivax gelatinosus LH2 complex, incorporated into a monoolein cubic phase.

The light-harvesting complex LH2 from a purple bacterium, Rubrivivax gelatinosus, has been incorporated into the Q230 cubic phase of monoolein. We measured the self-diffusion of LH2 in detergent solution and in the cubic phase by fluorescence recovery after photobleaching. We investigated also the absorption and fluorescence properties of this oligomeric membrane protein in the cubic phase, in comparison with its beta-octyl glucoside solution. In these experiments, native LH2 and LH2 labeled by a fluorescent marker were used. The results indicate that the inclusion of LH2 into the cubic phase induced modifications in the carotenoid and B800 binding sites. Despite these significant perturbations, the protein seems to keep an oligomeric structure. The relevance of these observations for the possible crystallization of this protein in the cubic phase is discussed.

Diffusion↗
PubMed2001

Dimer interface of transmembrane domains for neu/erbB-2 receptor dimerization and transforming activation: a model revealed by molecular dynamics simulations.

The specific point mutation Val-->Glu664 within the transmembrane domain of the neu/erbB-2 receptor is associated with increased receptor dimerization and increased receptor tyrosine kinase activity resulting in malignant transformation of cells. It is well established that Glu and residues in proximity are necessary for receptor dimerization but many studies suggest that other intramembrane constraints, not yet elucidated, are determinant for transformation. In this work, we investigated dimer models both to understand the structural role of the Glu mutation in the transmembrane domain association and to determine helix-helix contacts required for oncogenic transformation. Different types of helix-helix association based on data resulting from Cys mutational studies of the full wild receptor and spectroscopic data of transmembrane neu peptides have been explored by molecular dynamics simulations. The study leads to propose a model for the dimeric association of the transmembrane domains of the oncogenic neu receptor showing left-handed interactions of the two helices stabilized by symmetrical hydrogen bonding interactions involving the Glu side chain on one helix and the facing carbonyl of Ala661 on the second helix. Contacting residues observed in the symmetric interface explain the transforming activity or the non transforming activity of many neu mutants. Moreover the left-handed coiled coil structure is fully consistent with recent results proving the role of rotational linkage of the transmembrane domain with the kinase domain. Comparison between the predicted dimer model and those presumed from experiments strongly suggests helix flexibility in the extracellular juxtamembrane region.

Amino Acid Sequence↗
PubMed2000

Structure prediction of the dimeric neu/ErbB-2 transmembrane domain from multi-nanosecond molecular dynamics simulations.

Dimerization of the neu/ErbB-2 receptor tyrosine kinase is a necessary but not a sufficient step for signaling. Despite the efforts expended to identify the molecular interactions responsible for receptor-receptor contacts and particularly those involving the transmembrane domain, structural details are still unknown. In this work, molecular dynamics simulations of the helical transmembrane domain (TM) of neu and ErbB-2 receptors are used to predict their dimer structure both in the wild and oncogenic forms. A global conformational search method, applied to define the best orientations of parallel helices, showed an energetically favorable configuration with the specific mutation site within the interface, common for both the nontransforming and the transforming neu/ErbB-2 TM dimers. Starting from this configuration, a total of 10 simulations, about 1.4 ns each, performed in vacuum, without any constraints, show that the two helices preferentially wrap in left-handed interactions with a packing angle at about 20 degrees. The resulting structures are nonsymmetric and the hydrogen bond network analysis shows that helices experience pi local distortions that facilitate inter-helix hydrogen bond interactions and may result in a change in the helix packing, leading to a symmetric interface. For the mutated sequences, we show that the Glu side chain interacts directly with its cognate or with carbonyl groups of the facing backbone. We show that the connectivity between interfacial residues conforms to the knobs-into-holes packing mode of transmembrane helices. The dimeric interface described in our models is discussed with respect to mutagenesis studies.

Amino Acid Sequence↗
PubMed2000

Protein turnover modifications induced by the protein feeding pattern still persist after the end of the diets.

This study was undertaken to determine whether the protein feeding pattern could induce chronic adaptation of protein turnover. After a 15-day adaptive period, elderly (68 yr) and young (26 yr) women received, for 14 days, a diet providing 200 KJ x kg fat-free mass (FFM)(-1) x day(-1), where the daily protein intake (1.7 g protein x kg FFM(-1) x day(-1)) was either spread over 4 meals in the spread pattern or mainly (80%) consumed at noon in the pulse pattern. One day after the end of the dietary treatment, whole body leucine kinetics were measured by use of a continuous [(13)C]leucine infusion, both in the postabsorptive state and in the same fed state. The pulse pattern was able to induce, in young as in elderly women, a lower postabsorptive leucine oxidation and endogenous leucine flux than the spread pattern and improved the responsiveness of nonoxidative leucine disposal during 4-h oral feeding. Thus the pulse pattern was able to induce chronic regulation of protein metabolism in young as in elderly women.

Adaptation, Physiological↗
PubMed1999

Molecular dynamics simulations of the ErbB-2 transmembrane domain within an explicit membrane environment: comparison with vacuum simulations.

Two 500-ps molecular dynamics simulations performed on the single transmembrane domain of the ErbB-2 tyrosine kinase receptor immersed in a fully solvated dilauroylphosphatidyl-ethanolamine bilayer (DLPE) are compared to vacuum simulations. One membrane simulation shows that the initial alpha helix undergoes a local pi helix conversion in the peptide part embedded in the membrane core similar to that found in simulation vacuum. Lipid/water/peptide interaction analysis shows that in the helix core, the intramolecular peptide interactions are largely dominant compared to the interactions with water and lipids whereas the helix extremities are much more sensitive to these interactions at the membrane interfaces. Our results suggest that simulations in a lipid environment are required to understand the dynamics of transmembrane helices, but can be reasonably supplemented by in vacuo simulations to explore rapidly its conformational space and to describe the internal deformation of the hydrophobic core.

Amino Acid Sequence↗
PubMed1999

Systemic characteristics of chronic arthritis induced by transfer of human rheumatoid synovial membrane into SCID mice (human/murine SCID arthritis).

Erosive human/murine (hu/mu) SCID arthritis, caused by unilateral engrafting of human rheumatoid arthritis synovial membrane (RA-SM) in the knee joints of SCID mice, was monitored for up to 18 weeks by scintigraphic, radiological, morphological and immunohistochemical analyses.(99m)Tc-DPD scintigraphy and histology revealed secondary, oligoarticular spreading of arthritis to contralateral knees and hips, but not to forelimb joints. Also, there were no extraarticular manifestations. At 18 weeks, surviving human cells were found within the pannus, but not directly at the cartilage erosion front, where fibroblast-like cells and macrophages of murine origin predominated. The latter cells also predominated in secondarily affected joints, where no human cells were detectable. Preventive depletion of murine NK-cells by anti-asialo-GMI antibodies, to check the influence of NK cells independently of strain and MHC system, combined with application of autologous human PBMN cells, had virtually no effects on the disease process. The completeness of the SCID defect was not critical, i.e. T cells were completely absent in the organs examined, and the presence of a few B cells in the spleen did not correspond to particular disease features. The SCID defect itself had a clear impact, since, in the chronic phase, SCID.bg and RAG-2(-/-)knockout mice developed less consistent pathological/scintigraphic signs of disease than SCID mice. Thus, unilaterally-induced hu/mu SCID arthritis is an oligoarticular disorder of the hindlimbs. Murine macrophages and fibroblast-like cells appear responsible for tissue destruction in engrafted and non-engrafted arthritic joints.

Animals↗
PubMed1998

Time resolved fluorescence properties of phenylalanine in different environments. Comparison with molecular dynamics simulation.

Time resolved fluorescence of the phenylalanine residue (Phe) alone and included in the transmembrane domain (TMD) sequences of the epidermal growth factor receptor (EGFR) and ErbB-2 was studied using the synchrotron radiation source of light, and compared to molecular dynamics (MD) simulations. The fluorescence intensity decay is strongly sensitive to the environment. A mono-exponential decay was obtained for Phe amino acid alone in two different solvents and for Phe included in EGFR transmembrane sequence, with fluorescence lifetime values varying from 1.7 ns (EGFR) to 7.4 ns (Phe dissolved in water). In ErbB-2 transmembrane sequence three lifetimes were detected. The relative amplitude of the shortest one (0.14 ns) is smaller than 10%, whereas the others (0.6 and 2.2 ns) are almost equally represented. They have been attributed to different rotamers exchanging slowly. This interpretation is supported by MD simulations which evidence transitions in time series of the chi 1 dihedral angle of Phe observed in the case of ErbB-2. The anisotropy decays are similar for both peptides and indicate the presence of a correlation time in the nanosecond range (1-4 ns) and the probable existence of a very fast one (< 0.05 ns). Autocorrelation functions computed from MD simulations corroborate these results.

Amino Acid Sequence↗
PubMed1998

[Late potentials in chronic alcoholics].

OBJECTIVE: Cardiac arrest is the most frequent cause of death in chronic alcoholics. Detection of late potentials in this population could be helpful in screening from early signs of myocardial disorders and identifying patients at risk of severe ventricular dysrythmia. PATIENTS AND METHODS: A prospective study of late potentials was conducted in 53 subjects (mean age 49 +/- 10 years) with a history of long-standing alcohol abuse (mean 13.6 +/- 8.5 years, mean daily alcohol intake 86 +/- 30 g). After a period of abstinence, the following explorations were performed: liver tests, liver biopsy, electrocardiogram, echocardiography, Holter recording. RESULTS: Among the 53 patients, 37% were positive for 2 of the 3 criteria for late potentials. There was a strong correlation between the duration of alcohol abuse and presence of late potentials (p = 0.006, r = 0.37). The percentage of hepatic steatosis was higher in alcoholic subjects with late potentials (34% versus 23%; p = 0.05) and was correlated with the number of positive criteria for late potentials (p = 0.05, r = 0.328). Finally, the presence of late potentials was also correlated with the following laboratory results: serum gamma glutamyltranspeptidase (p = 0.031), serum aspartate amino transferase (p = 0.033), serum alkaline phosphatases (p = 0.0025). CONCLUSION: Late potentials can be detected easily although their prognostic value remains to be determined. They could be an early marker of infraclinical myocardial lesions.

Adult↗
PubMed1998

[Signal averaging electrocardiography in chronic alcoholism].

Cardiovascular death is the main cause of mortality in chronic alcoholics, perhaps due to a pro-arrhythmogenic effect of alcohol associated with infraclinical myocardial lesions. The authors investigated prospectively 41 patients (average age: 49.7 years) who were chronic alcoholics but had no acute alcoholic episodes for cardiac disease (ECG, signal averaging for late ventricular potentials, echocardiography and Holter ECG monitoring) and hepatic disease (liver biopsy). The history of alcoholism was 14 +/- 9 years, the quantity of alcohol ingested before they stopped drinking being 89 +/- 31 grammes/day. Thirty per cent of patients displayed 2 or 3 criteria of late ventricular potentials (LP). The authors demonstrated a correlation between the daily quantity of alcohol consumed before stopping drinking and the duration of the filtered QRS complex (p = 0.02). Moreover, the frequency of fatty infiltration found on liver biopsy, greater in alcoholics with LP (35% versus 19%, p = 0.025) correlated with the amplitude of the last 40 ms of the average QRS (p = 0.0485), with the duration of potentials of less than 40 microvolts (p = 0.05) and, above all, with the number of criteria of LP (p = 0.02). Finally, the presence of LP was also related to the following biological abnormalities: GGT (p = 0.027), ASAT (p = 0.046), ALAT (p = 0.039). The ECG abnormalities may reflect early infra-clinical myocardial lesions secondary to cellular metabolic abnormalities perhaps analogous to the fatty hepatic changes. However, the prognostic value of these signal-averaging ECG abnormalities remains unknown.

Adult↗
PubMed1997

Molecular modeling of c-erbB2 receptor dimerization: coiled-coil structure of wild and oncogenic transmembrane domains--stabilization by interhelical hydrogen bonds in the oncogenic form.

Dimerization models of c-erbB2 transmembrane domains (Leu651-Ile675) are studied by molecular mechanics and molecular dynamics simulations. Both wild and Glu mutated transmembrane helices exhibit the same relative orientation for favorable associations and dimerize preferentially in left-handed coiled-coil structures. The mutation point 659 belongs to the interfacing residues, and in the transforming domain, symmetric hydrogen bonds between Glu carboxylic groups stabilize the dimeric structure. The same helix packing found for the wild dimers, except side-chain-side-chain hydrogen bonds, suggests that the transmembrane domains dimerize according to similar process. Structural and energetical characterization of the models are presented.

Amino Acid Sequence↗
PubMed1997

Insight into signal transduction: structural alterations in transmembrane helices probed by multi-1 ns molecular dynamics simulations.

The hypothesis of structural alteration in transmembrane helices for signal transduction process is viewed by molecular dynamics simulation techniques. For the c-erbB-2 transmembrane domain involved in oncogenicity, the occurrence of conformational changes has been previously described as transition from the alpha to pi helix. This dynamical feature is thoroughly analyzed for the wild phenotype and oncogenic sequences from a series of 18 simulations carried out on one nanosecond time scale. We show that these structural events do not depend upon the conditions of simulations like force field or starting helix coordinates. We demonstrate that the oncogenic mutations Val659 Glu, Gln and Asp do not prevent the transition. Furthermore, we show that beta branched residues, in conjunction with Gly residues in the c-erbB-2 sequence, act as destabilizers for the alpha helix structure, pi deformations are tightly related to other local structural motifs found in soluble and membrane proteins. These structural alterations are discussed in term of structure-activity relationships for the c-erbB-2 activating mechanism mediated by transmembrane domain dimerization.

Amino Acid Sequence↗
PubMed1997

Late ventricular potentials and heavy drinking.

OBJECTIVES: To assess the effects of chronic drinking on detection of low amplitude signals, and to determine the relation between late ventricular potentials (LVP) and liver biopsy findings. DESIGN: Prospective study. SETTING: General hospital. PATIENTS: 41 consecutive chronic alcoholics without known pre-existing heart disease. METHODS: About four days after each patient's last alcoholic drink, ECG, echocardiography, signal averaged electrocardiogram, liver biopsy, and blood tests were performed. RESULTS: Twenty eight per cent of patients had evidence of LVP. There was a correlation between the percentage of steatosis of the hepatic biopsy and the amplitude of the last 40 ms of average QRS (P = 0.04), the duration of the terminal low amplitude QRS signal (P = 0.05), and the number of positive criteria of late potentials (P = 0.02). CONCLUSIONS: Chronic drinking sufficient to cause steatosis is associated with positive findings on the signal averaged ECG.

Alcoholism↗
PubMed1996

Correlated motions and propagation of the effect of a local conformational change in the transmembrane helix of the c-erb B2 encoded protein and its V659E mutant, studied by molecular dynamics simulations.

A detailed study of the dynamical behavior of the 29-residue peptide including the transmembrane domain of p185c-erbB2 oncogene-encoded protein and of its V659E mutant is presented. In a first part of this work we analyse equal time correlation coefficients between the backbone dihedral angle fluctuations. Concerted motions are observed in the wild type transmembrane alpha-helix but not in the corresponding V659E intramembrane domain. The difference observed in the correlation pattern is attributed to the single amino acid replacement. In a second part, we investigate the propagation of the effect of a local conformational change along the transmembrane segment, one of the dominant hypotheses for signal transduction mechanisms of transmembrane receptors. The analysis of angular time correlation functions together with that of the response of the different residues to a local disturbance applied at the N-terminal side evidences a propagation phenomenon for the wild type peptide. This effect is much less clear for the mutated peptide. Furthermore we show that the first one is much more flexible than the second one.

Cell Membrane↗
PubMed1996

Orthotopic implantation of inflamed synovial tissue from RA patients induces a characteristic arthritis in immunodeficient (SCID) mice.

The objective of this work was to study in more detail the human/murine SCID arthritis model with special emphasis on characteristic features initiated by rheumatoid arthritis (RA) synovial membrane (SM) as compared to appropriate control tissues. Small tissue samples from RA-SM, healthy lymph node, healthy SM, and granulomatous tissue of human origin were implanted into the left knee joint of mice with severe combined immunodeficiency (SCID), and the joints were analysed histologically after 7 days. In addition, a time course study, including non-invasive monitoring by serological parameters (human IgM, IgG, and IL-6) and Tc-99m-scintigraphy, was performed for up to 4 weeks on RA-SM recipients. All tissue implants induced transient exudative joint inflammation while RA-SM initiated a characteristic arthritis with pannus tissue of high cellular density, erosion, multinuclear giant cells, lining cell hyperplasia, fibroblast-like cell layers, chondroideal metaplasia, and fibrin deposits. Significantly elevated levels of human immunoglobulin and characteristic signs of chronic inflammation persisted for more than 4 weeks. We conclude that the hu/mu SCID arthritis with RA-SM implants comprises features of non-specific inflammation which is also transiently seen with control tissues but develops characteristic features of chronic RA-like synovitis thereafter.

Animals↗
PubMed1996

Detailed description of an alpha helix-->pi bulge transition detected by molecular dynamics simulations of the p185c-erbB2 V659G transmembrane domain.

Molecular dynamics simulations of a 29-residue peptide including the transmembrane domain of the V659G mutant of the c-erbB2 protein demonstrate important dynamical behavior. Although the alpha helix is the structure commonly assumed for transmembrane hydrophobic segments, we found that hydrogen bond rearrangements can occur, giving rise to a structural deformation termed pi bulge stabilized by successive hydrogen bonds of pi helix type. A series of simulations enables us to give a detailed description, at the atomic level, of the alpha helix->pi bulge transition. The major consequence of this deformation covering one and a half turn of helix results in a noticeable shift around the helix axis of the C-Terminal residues relatively to those of the N-terminus. Such a deformation closely related to structural motifs described in the literature, induces a change in the distribution of the residues along the helix faces which could modulate the protein activity mediated by a dimerization process.

Amino Acid Sequence↗