Biomedical subjects

B P Roques

Publications and source records attributed to B P Roques.

At least 19 recordsLinked to original sources

PubMed2006

Interaction between the HIV-1 protein Vpr and the adenine nucleotide translocator.

The HIV-1 protein Vpr circulates in the serum of seropositive individuals and in the cerebrospinal fluid of AIDS patients with neurological disorders. Vpr triggers apoptosis of numerous cell types after extracellular addition, vpr gene transfer or in the context of viral infection. Moreover, in vivo, transgenic mice over-expressing Vpr have enhanced T lymphocytes apoptosis. In previous studies, we suggested that the Vpr apoptotic activities were because of its binding to the adenine nucleotide translocator (ANT), a mitochondrial ATP/ADP antiporter. To specify this interaction, fragments of both proteins were synthesized and used in biochemical and biophysical experiments. We demonstrate here that in vitro, the (27-51) and (71-82) Vpr peptides bind to a region encompassing the first ANT intermembrane space loop and part of its second and third transmembrane helices. Computational analysis using a docking program associated to dynamic simulations enabled us to construct a three-dimensional model of the Vpr-ANT complex. In this model, the N-terminus of Vpr plunges in the ANT cavity whereas the Vpr C-terminal extremity is located at the surface of the ANT allowing possible interactions with a third partner. These results could be used to design molecules acting as pro-apoptotic Vpr analogs or as apoptosis inhibitors preventing the Vpr-ANT interaction.

Animals↗

PubMed2005

Facilitation of enkephalins-induced delta-opioid behavioral responses by chronic amisulpride treatment.

The endogenous opioid system is known to have a great influence on the dopaminergic system. Conversely, blockade of the dopaminergic system in D2 receptor knock-out mice triggers an increase in enkephalin supporting the important physiological relationship between both systems. Therefore, the aim of this study was to investigate whether or not chronic treatment with the specific D2 antagonist amisulpride (20mg/kg, i.p., twice daily for 5 days) could lead to a facilitation of behavioral effects of enkephalins, protected from their enzymatic degradation by the dual inhibitor N-[(R,S)-2-benzyl-3[(S)(2-amino-4-methylthio)butyl dithio]-1-oxopropyl]-l-phenylalanine benzyl ester (RB101) (5mg/kg, i.v.) in mice. RB101 induced an increase in locomotor activity, antidepressant-like effects in the forced swim test, and antinociceptive effects in the hot-plate test. Chronic treatment with amisulpride potentiated the action of RB101 and this effect seemed to be restricted to behavioral responses induced by opioids acting on delta-opioid receptors (locomotor activity and forced swim test). This was confirmed by the use of the selective delta-opioid receptor agonist, (+)-4-[alpha-R*)-alpha-((2S*,5R*)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80; 2.5mg/kg, i.p.), and antagonist, naltrindole (5mg/kg, i.p.). Considering the involvement of delta-opioid receptors in mood regulation, the interaction between amisulpride and RB101 could lead to a new therapeutic approach in the treatment of some mood disorders.

Amisulpride↗

PubMed2005

Physiological control of emotion-related behaviors by endogenous enkephalins involves essentially the delta opioid receptors.

The endogenous pentapeptide enkephalins bind to the mu and delta opioid receptors, with a slightly higher affinity for the latter. It remains a controversy regarding the respective physiological role of mu and delta opioid receptors in the control of emotion and motivation. One of the difficulties to investigate this problem is the low tonic extracellular release of enkephalins in various brain structures. To overcome this problem the synaptic levels of these pentapeptides were enhanced by inhibition of enzymes involved in their catabolism with the selective inhibitor H3N-CH(CH2-CH2-S-CH3)-CH2-S-S-CH2-CH(CH2phi)-CONH-CH(CH2phi)-COOCH2phi (RB101). This compound was shown to increase the extracellular levels and lifetime of endogenous enkephalins. Similar responses were obtained in wild-type and mu opioid receptor knockout mice following RB 101 administration in behavioral tests measuring locomotor activity, anxiety (elevated O-maze), and motivation (forced swim test and conditioned suppression of motility). In contrast, RB 101 led to antinociceptive responses only in wild-type animals using hot plate and tail immersion tests. These results clearly demonstrate the critical role of delta opioid receptors activated by the endogenous opioid peptides, in the physiological control of emotion- and motivation-related behaviors. In contrast, antinociceptive modulation, at least with respect to thermal nociceptive stimuli, involves enkephalin-activated mu opioid receptors. These findings could open new perspectives in the treatment of mood disorders using either inhibitors of enkephalin catabolism or delta opioid agonists.

Analysis of Variance↗

PubMed2004

Target specificity of human immunodeficiency virus type 1 NCp7 requires an intact conformation of its CCHC N-terminal zinc finger.

The modification of zinc-binding residues inside the conserved CCHC motif of human immunodeficiency virus type 1 NCp7, in particular into CCHH, induces a complete loss of infectivity. Since the mutant His28NCp7 has been shown to be devoid of infectivity in vivo, the structure-function relationships of the mutant His28(12-53)NCp7 were investigated by nuclear magnetic resonance and surface plasmonic resonance. Although the Cys28-->His mutation modifies drastically the structure of the core domain (residues 12 to 53) of NCp7, His28(12-53)NCp7 still interacts with a 10-fold-lower affinity to specific nucleic acid targets, such as SL3, a stem-loop critically involved in viral RNA packaging, and without affinity change with the nonspecific, single-stranded nucleic acid poly(T). Moreover, His28(12-53)NCp7 and native (12-53)NCp7 displayed the same affinity with reverse transcriptase, but the natures of the complexes are probably different, accounting for the drastic reduction in the amount of RNA packaged in the mutated virus. We propose a structural model of His28(12-53)NCp7 that provides insights into the NCp7 structural features necessary for target recognition and that shows that the specific native structure of the zinc finger domain is strictly required for the optimal target selectivity of NCp7.

Amino Acid Motifs↗

PubMed2003

NMR structure of the HIV-1 regulatory protein VPR.

The human immunodeficiency virus type 1 (HIV-1) genome encodes a highly conserved regulatory gene product, Vpr (96 residues, 14kDa), which is incorporated into virions. In the infected cells, Vpr, expressed late in the virus cycle, is believed to function in the early phases of HIV-1 replication, such as nuclear migration of pre-integration complex, transcription of the proviral genome, viral multiplication by blocking cells in G2 phase and regulation of apoptosis phenomenon. Vpr has a critical role in long term AIDS disease by inducing infection in non-dividing cells such as monocytes and macrophages. To gain insight into the structure-function relationships of Vpr, the (1-96)Vpr protein was synthesized with 22 labeled amino acids. Its 3D structure was analyzed in the presence of CD(3)CN and in pure water at low pH and refined by restrained simulated annealing. The structure of the protein is characterized by three well-defined alpha-helices: 17-33, 38-50 and 56-77 surrounded by flexible N and C-terminal domains. In contrast to the structure obtained in the presence of TFE, the three alpha-helices are folded around a hydrophobic core constituted of Leu, Ile, Val and aromatic residues as illustrated by numerous long range NOEs. This structure accounts for the interaction of Vpr with different targets.

Amino Acid Sequence↗

PubMed2003

Involvement of brain endogenous cholecystokinin in stress-induced impairment of spatial recognition memory.

The central fragment of cholecystokinin, CCK8, plays a critical role in stress-related changes in behavior and memory. Therefore, we investigated whether the endogenous cholecystokininergic system is involved in the impairment of attention and/or memory induced by stressful conditions. Plasma corticosterone concentrations increased three-fold and plasma adrenocorticotropin (ACTH); concentrations increased five-fold when rats were maintained in the open arm of an elevated plus maze for 5 min. The same stress conditions impaired spatial recognition in the two-trial memory task. In addition, this stress led to a significant decrease in the extracellular levels of cholecystokinin-like immunoreactivity in the dorsal subiculum/CA1 of the hippocampus and partially suppressed the increase obtained during the acquisition phase of memory. This suggests that the cholecystokininergic system in the hippocampus is involved in stress-induced impairment of spatial recognition memory.

Adrenocorticotropic Hormone↗

PubMed2003

[Plusridisciplinary approaches to understand the neutoxic effects of Ecstasy].

Recreational use of the illicit drug ecstasy (MDMA) has increased dramatically in recent years. Although generally regarded as relatively safe, several studies have suggested a toxic effect on brain neurons in animals and possibly in humans. Others have demonstrated that the neurotoxic effects of MDMA on 5-HT neurons may be reversible, while the effects of MDMA on memory function may be long-lasting. To clearly evaluated the risks of repeated MDMA use, it is crucial to characterize its targets. Moreover, in animal, the direct injection of MDMA into the brain does not reproduce neurotoxic effects observed after peripheral administration. This fact suggests a possible implication of MDMA metabolites in the development of neurotoxicity, which should be identified. These metabolites may play a role in the long term psychological effects of MDMA. To be able to answer all, or at least most of the questions on the risks induced by repeated use of MDMA, a wide range of disciplines should be used, chemistry, genomic, proteomic and pharmacology.

Animals↗

PubMed2002

Zn(2+) binding properties of single-point mutants of the C-terminal zinc finger of the HIV-1 nucleocapsid protein: evidence of a critical role of cysteine 49 in Zn(2+) dissociation.

The two highly conserved Zn(2+) finger motifs of the HIV-1 nucleocapsid protein, NCp7, strongly bind Zn(2+) through coordination of one His and three Cys residues. To further analyze the role of these residues, we investigated the Zn(2+) binding and acid-base properties of four single-point mutants of a short peptide corresponding to the distal finger motif of NCp7. In each mutant, one Zn(2+)-coordinating residue is substituted with a noncoordinating one. Using the spectroscopic properties of Co(2+), we first establish that the four mutants retain their ability to bind a metal cation through a four- or five-coordinate geometry with the vacant ligand position(s) presumably occupied by water molecule(s). Moreover, the pK(a) values of the three Cys residues of the mutant apopeptide where His44 is substituted with Ala are found by (1)H NMR to be similar to those of the native peptide, suggesting that the mutations do not affect the acid-base properties of the Zn(2+)-coordinating residues. The binding of Zn(2+) was monitored by using the fluorescence of Trp37 as an intrinsic probe. At pH 7.5, the apparent Zn(2+) binding constants (between 1.6 x 10(8) and 1.3 x 10(10) M(-)(1)) of the four mutants are strongly reduced compared to those of the native peptide but are similar to those of various host Zn(2+) binding proteins. As a consequence, the loss of viral infectivity following the mutation of one Zn(2+)-coordinating residue in vivo may not be related to the total loss of Zn(2+) binding. The pH dependence of Zn(2+) binding indicates that the coordinating residues bind Zn(2+) stepwise and that the free energy provided by the binding of a given residue may be modulated by the entropic contribution of the residues already bound to Zn(2+). Finally, the pK(a) of Cys49 in the holopeptide is found to be 5.0, a value that is at least 0.7 unit higher than those for the other Zn(2+)-coordinating residues. This implies that Cys49 may act as a switch for Zn(2+) dissociation in the distal finger motif of NCp7, a feature that may contribute to the high susceptibility of Cys49 to electrophilic attack.

Alanine↗

PubMed2002

NMR structure of the HIV-1 regulatory protein Vpr in H2O/trifluoroethanol. Comparison with the Vpr N-terminal (1-51) and C-terminal (52-96) domains.

The human immunodeficiency virus type 1, HIV-1, genome encodes a highly conserved regulatory gene product, Vpr (96 amino acids), which is incorporated into virions in quantities equivalent to those of the viral Gag protein. In infected cells, Vpr is believed to function during the early stages of HIV-1 replication (such as transcription of the proviral genome and migration of preintegration nuclear complex), blocks cells in G2 phase and triggers apoptosis. Vpr also plays a critical role in long-term AIDS disease by inducing viral infection in nondividing cells such as monocytes and macrophages. To gain deeper insight of the structure-function relationship of Vpr, the intact protein (residues 1-96) was synthesized. Its three-dimensional structure was analysed using circular dichroism and two-dimensional 1H- and 15N-NMR and refined by restrained molecular dynamics. In addition, 15N relaxation parameters (T1, T2) and heteronuclear 1H-15N NOEs were measured. The structure of the protein is characterized by a well-defined gamma turn(14-16)-alpha helix(17-33)-turn(34-36), followed by a alpha helix(40-48)-loop(49-54)-alpha helix(55-83) domain and ends with a very flexible C-terminal sequence. This structural determination of the whole intact Vpr molecule provide insights into the biological role played by this protein during the virus life cycle, as such amphipathic helices are believed to be involved in protein-lipid bilayers, protein-protein and/or protein-nucleic acid interactions.

Amino Acid Sequence↗

PubMed2001

HIV-1 nucleocapsid protein and the secondary structure of the binary complex formed between tRNA(Lys.3) and viral RNA template play different roles during initiation of (-) strand DNA reverse transcription.

In human immunodeficiency virus type 1 (HIV-1), the tRNA(Lys.3) primer and viral RNA template can form a specific complex that is characterized by extensive inter- and intramolecular interactions. Initiation of reverse transcription from this complex has been shown to be distinguished from subsequent elongation by early pausing events, such as at the +1 and +3 nucleotide positions. One major concern regarding the biological relevance of these results is that most kinetic studies of HIV-1 reverse transcription have been performed using tRNA(Lys.3)-viral (v) RNA complexes that were formed by heat annealing. In contrast, tRNA(Lys.3) in viruses is placed onto the primer binding site by nucleocapsid (NC) sequences of the Gag protein. In this study, we have further characterized the initiation features of reverse transcription in the presence of HIV-1 NC protein. In contrast to results obtained with a heat-annealed tRNA(Lys.3).vRNA complex, we found that polymerization reactions catalyzed by HIV-1 reverse transcriptase did not commonly pause at the +1 nucleotide position when a NC-annealed RNA complex was used, and that this was true regardless whether NC was actually still present during reverse transcription. This activity of NC required both zinc finger motifs, as demonstrated by experiments that employed zinc finger-mutated forms of NC protein (H23C NC and ddNC), supporting the involvement of the zinc fingers in the RNA chaperone activity of NC. However, NC was not able to help reverse transcriptase to escape the +3 pausing event. Mutagenesis of a stem structure within the tRNA(Lys.3). vRNA complex led to disappearance of the +3 pausing event as well as to significantly reduced rates of reverse transcription. Thus, this stem structure is essential for optimal reverse transcription, despite its role in promotion of the +3 pausing event.

Amino Acid Motifs↗

PubMed2001

Long lasting antinociceptive properties of enkephalin degrading enzyme (NEP and APN) inhibitor prodrugs.

Prodrugs of phosphinic dual inhibitors of the enkephalin degrading enzymes, neutral endopeptidase (NEP) and aminopeptidase N (APN), corresponding to the formula H(3)N(+)CH(R(1))P(O)(OR)CH(2)CH(CH(2)Bip)CONHCH(CH(3))COOCH(2)Ph, with R(1) = CH(3) or Ph and R being a benzyl ester, a S-acyl-2-thioethyl derivative, or an acyloxyalkyl group, were synthesized to improve the poor central bioavailability of their precursors. As expected, these compounds (50 mg/kg, iv or ip) induced long lasting ( approximately 2 h) antinociceptive responses in the hot plate test in mice with a ceiling effect varying between 25 and 42% of analgesia. A very rapid hydrolysis of the carboxylate ester contrasting with a slow deprotection of the phosphinate group (t(1/2) approximately 1 h) was observed in serum while 80% of free drug was obtained after 1 h incubation with brain membranes. These results account for the long duration of action observed with these prodrugs.

Aminopeptidases↗

PubMed2001

Determination of the pK(a) of the four Zn2+-coordinating residues of the distal finger motif of the HIV-1 nucleocapsid protein: consequences on the binding of Zn2+.

The nucleocapsid protein NCp7 of human immunodeficiency virus type 1 is characterized by two highly conserved CCHC motifs that bind Zn2+ strongly. To elucidate the striking pH-dependence of the apparent Zn2+-binding constants of these motifs further, we investigated, using 1H NMR, potentiometry and fluorescence spectroscopy, the acid-base properties of the four Zn2+-coordinating residues of (35-50)NCp7, a peptide corresponding to the distal finger motif of NCp7. With the exception of the H(beta2) proton of Cys39, the pH-dependence of the H(beta) proton resonances of the three Cys residues and, the H(delta) and H(epsilon) resonances of His44 in the apopeptide could be fitted adequately with a single pK(a). This suggests that the protonating groups are non-interacting, a feature that was confirmed by a potentiometric titration. The pK(a) of His44, Cys36, Cys39, and Cys49 in the apopeptide were found to be 6.4, 8.0, 8.8 and 9.3, respectively. Accordingly, the deprotonation is almost sequential and may thus induce a sequential binding of Zn2+ to the four coordinating residues. The high pK(a) of Cys49 is probably related to the negative charge of the neighboring Asp48. Such a high pK(a) may be a general feature in nucleocapsid proteins (NCs), since an acidic residue generally occupies the (i-1) position of the C-terminal Cys residue of single-finger NCs and distal finger motifs in two-finger NCs. Molecular dynamics simulation suggested the formation of a hydrogen bonded network that weakly structured the Cys36-Cys39 segment in the apopeptide. This network depends on the protonation state of Cys36 and may thus explain the biphasic behavior of the pH-dependence of the Cys39 H(beta2) resonance. Finally, the pK(a) values were used to build up a model describing the coordination of Zn2+ to (35-50)NCp7 at equilibrium. It appears that each protonation step of the coordination complex decreases the Zn2+-binding constant by about four orders of magnitude and that a significant dissociation of Zn2+ from the holopeptide can be achieved in acidic cell compartments.

Amino Acid Sequence↗

PubMed2001

Mutant mice lacking the cholecystokinin2 receptor show a dopamine-dependent hyperactivity and a behavioral sensitization to morphine.

Cholecystokinin2 (CCK2) receptor-deficient mice were used to analyze the in vivo function of CCK2 receptor and especially the incidence of this gene invalidation on enkephalinergic and dopaminergic systems. Hyperlocomotor activity of CCK2 receptor-deficient mice was suppressed by a selective D2 antagonist but not by a D1 antagonist. Injection of amphetamine induced a hyperlocomotor activity in both groups of mice while mutant mice were less sensitive to cocaine. Administration of 6 mg/kg of morphine once every 2 days for 5 days significantly (P<0.05) enhanced motor activity the last day compared to the first day, only in CCK2 receptor-deficient mice. These results emphasize the role of CCK2 receptors in counteracting the effects of dopaminergic systems and suggest that CCK2 receptor invalidation could lead to a slight behavioral sensitization.

Analgesics, Opioid↗

PubMed2001

Highly sensitive and selective fluorescence assays for rapid screening of endothelin-converting enzyme inhibitors.

The highly potent vasoconstrictor peptide endothelin (ET) is generated from an inactive precursor, big endothelin (bET), by endothelin-converting enzyme (ECE). ECE is a phosphoramidon-sensitive zinc metallopeptidase, which is closely related to neprilysin (neutral endopeptidase). It is possible that compounds which inhibit the formation of ET may be used as new drugs for the treatment of cardiovascular diseases. Such an approach requires a fast, simple and selective assay to measure ECE activity, allowing rapid screening of inhibitors. We describe here two new ECE substrates based on the concept of 'intramolecularly quenched fluorescence' which may fulfill this aim. One, S(1) [Pya(21)-Nop(22)-bET-1(19--35)], is the (19--35) fragment of the natural peptide big-ET-1(1--38), which is modified by introducing the fluorescent amino acid, pyrenylalanine (Pya), in position 21 and a quencher, p-nitrophenylalanine (Nop), in position 22. The second substrate (S(2)) is a small peptide, Ac-Ser-Gly-Pya-Lys-Ala-Phe-Ala-Nop-Gly-Lys-NH(2), from a biased substrate peptide library. The recombinant, hECE-1c, cleaved both Pya(21)-Nop(22)-bET-1(19--35) and the natural substrate selectively between residues 21 and 22, whereas cleavage occurred between alanine and phenylalanine in the small peptide. In both cases, this generated intense fluorescence emission. The synthesis and kinetic parameters of these substrates are described. These assays, which can be used directly on tissue homogenates, are the most sensitive and selective described to date for ECE, and are easily automated for a high-throughput screening of inhibitors.

Amino Acid Sequence↗

PubMed2001

High-throughput fluorogenic assay for determination of botulinum type B neurotoxin protease activity.

Botulinum neurotoxins are responsible for botulism, a flaccid muscular paralysis caused by inhibition of acetylcholine release at the neuromuscular junction. This occurs by cleavage of conserved proteins involved in exocytosis such as synaptobrevin by the zinc metallopeptidase activity of the light chain of some botulinum neurotoxins. Botulism, for which there is presently no therapy available, is a relatively widespread disease that may result in death. Consequently, the development of drugs able to inhibit the hydrolytic activity of these neurotoxins is of great interest. Design and screening of such inhibitors could be largely facilitated by using high-throughput assays. With this aim, a novel in vitro test for quantifying the proteolytic activity of botulinum type B neurotoxin was developed. The substrate is the 60--94 fragment of human synaptobrevin-1 which was modified by introduction of the fluorescent amino acid l-pyrenylalanine in position 74 and a p-nitrophenylalanyl residue as quenching group in position 77. The cleavage of Syb 60-94 [Pya(74), Nop(77)] by the toxin active chain occurs selectively between residues 76 and 77 as in the case of the unmodified synaptobrevin and is directly quantified by measuring the strong fluorescence of the formed metabolite Syb 60-76 [Pya(74)]. This is the easiest, quickest, and cheapest assay described to date for measuring the proteolytic activity of botulinum type B neurotoxin. It can be easily automated for high-throughput screening. Moreover, amounts of about 3.5 pg/ml of botulinum type B neurotoxin could be detected by this method.

Biological Assay↗

PubMed2001

Heteronuclear NMR studies of the interaction of tRNA(Lys)3 with HIV-1 nucleocapsid protein.

Reverse transcription of HIV-1 viral RNA uses human tRNA(Lys)3 as a primer. Recombinant tRNA(Lys)3 was previously overexpressed in Escherichia coli, 15N-labelled and purified for NMR studies. It was shown to be functional for priming of HIV-1 reverse transcription. Using heteronuclear 2D and 3D NMR, we have been able to assign almost all the imino groups in the helical regions and involved in the tertiary base interactions of tRNA(Lys)3. This crucial step enabled us to address the question of the annealing mechanism of tRNA(Lys)3 by the nucleocapsid protein (NC) using heteronuclear NMR. Moreover, structural aspects of the tRNA(Lys)3/(12-53)NCp7 interaction have been characterised. The (12-53)NCp7 protein binds preferentially to the inside of the L-shape of the tRNA(Lys)3, on the acceptor and D stems, and at the level of the tertiary interactions between the D and T-psi-C loops. (12-53)NCp7 binding does not induce the melting of any single base-pair or unwinding of the tRNA(Lys)3 helical domains. Moreover, NMR provides a unique means to investigate the base-pairs that are destabilised by (12-53)NCp7 binding. Indeed, the measurements of the longitudinal relaxation time T1 and of the exchange time of the imino protons revealed two major regions sensitive to catalysis by the protein, namely the G6-U67 and T54(A58) pairs. It is interesting that for the biological role of the NC protein, these pairs could be the starting points of the tRNA melting required for the hybridisation to the viral RNA.

Anticodon↗

PubMed2001

Control of mitochondrial membrane permeabilization by adenine nucleotide translocator interacting with HIV-1 viral protein rR and Bcl-2.

Viral protein R (Vpr), an apoptogenic accessory protein encoded by HIV-1, induces mitochondrial membrane permeabilization (MMP) via a specific interaction with the permeability transition pore complex, which comprises the voltage-dependent anion channel (VDAC) in the outer membrane (OM) and the adenine nucleotide translocator (ANT) in the inner membrane. Here, we demonstrate that a synthetic Vpr-derived peptide (Vpr52-96) specifically binds to the intermembrane face of the ANT with an affinity in the nanomolar range. Taking advantage of this specific interaction, we determined the role of ANT in the control of MMP. In planar lipid bilayers, Vpr52-96 and purified ANT cooperatively form large conductance channels. This cooperative channel formation relies on a direct protein-protein interaction since it is abolished by the addition of a peptide corresponding to the Vpr binding site of ANT. When added to isolated mitochondria, Vpr52-96 uncouples the respiratory chain and induces a rapid inner MMP to protons and NADH. This inner MMP precedes outer MMP to cytochrome c. Vpr52-96-induced matrix swelling and inner MMP both are prevented by preincubation of purified mitochondria with recombinant Bcl-2 protein. In contrast to König's polyanion (PA10), a specific inhibitor of the VDAC, Bcl-2 fails to prevent Vpr52-96 from crossing the mitochondrial OM. Rather, Bcl-2 reduces the ANT-Vpr interaction, as determined by affinity purification and plasmon resonance studies. Concomitantly, Bcl-2 suppresses channel formation by the ANT-Vpr complex in synthetic membranes. In conclusion, both Vpr and Bcl-2 modulate MMP through a direct interaction with ANT.

Amino Acid Sequence↗

PubMed2001

Effects of the potent analgesic enkephalin-catabolizing enzyme inhibitors RB101 and kelatorphan on respiration.

We investigated whether the enkephalin-catabolizing enzyme inhibitors RB101 and kelatorphan, which have been shown to be potent analgesics, depress respiration as do opioid analgesics. Ventilation was measured in cats and rodents by the barometric method, in the awake state and during anesthesia. Tissue distribution of the inhibitors was either generalized (RB101, 40-160 mg/kg i.p.), largely restricted by the blood-brain barrier to the periphery (kelatorphan, 0.7-20 mg/kg i.v.), or restricted to the brainstem (i.c.v. injection of RB101 in the fourth ventricle). RB101 did not affect ventilation in any condition tested, and large doses of kelatorphan produced a naloxone-reversible increase in ventilation and breathing frequency. Thus endogenous opioids released during conditions of normal ventilation do not exert any depressant neuromodulatory effect on this function, even when their extracellular concentrations are increased by peptidase inhibitors. The differential effect of these inhibitors on ventilation and nociception is discussed. We conclude that kelatorphan and RB101 are devoid of respiratory-depressant effects and might be interesting pharmacological alternatives to morphine and other opioid agonists.

Analgesics↗