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Biomedical subjects

L Pardo

Publications and source records attributed to L Pardo.

At least 37 records · Page 2Linked to original sources
PubMed1998

Experimental and theoretical characterization of the high-affinity cation-binding site of the purple membrane.

Binding of Mn2+ or Mg2+ to the high-affinity site of the purple membrane from Halobacterium salinarium has been studied by superconducting quantum interference device magnetometry or by ab initio quantum mechanical calculations, respectively. The binding of Mn2+ cation, in a low-spin state, to the high-affinity site occurs through a major octahedral local symmetry character with a minor rhombic distortion and a coordination number of six. A molecular model of this binding site in the Schiff base vicinity is proposed. In this model, a Mg2+ cation interacts with one oxygen atom of the side chain of Asp85, with both oxygen atoms of Asp212 and with three water molecules. One of these water molecules is hydrogen bonded to both the nitrogen of the protonated Schiff base and the Asp85 oxygen. It could serve as a shuttle for the Schiff base proton to move to Asp85 in the L-M transition.

Aspartic Acid↗
PubMed1998

Selective binding of the TATA box-binding protein to the TATA box-containing promoter: analysis of structural and energetic factors.

We report the results of an energy-based exploration of the components of selective recognition of the TATA box-binding protein (TBP) to a TATA box sequence that includes 1) the interaction between the hydrophobic Leu, Pro, and Phe residues of TBP with the TA, AT, AA, TT, and CG steps, by ab initio quantum mechanical calculations; and 2) the free energy penalty, calculated from molecular dynamics/potential of mean force simulations, for the conformational transition from A-DNA and B-DNA into the TA-DNA form of DNA observed in a complex with TBP. The GTAT, GATT, GAAT, and GTTT tetramers were explored. The results show that 1) the discrimination of TA, AT, AA, TT, or CG steps by TBP cannot rest on their interaction with the inserting Phe side chains; 2) the steric clash between the bulky and hydrophobic Pro and Leu residues and the protruding -NH2 group of guanine is responsible for the observed selectivity against any Gua-containing basepair; 3) the Pro and Leu residues cannot selectively discriminate among TA, AT, AA, or TT steps; and 4) the calculated energy required to achieve the TA-DNA conformation of DNA that is observed in the complex with TBP appears to be a key determinant for the observed selectivity against the AT, AA, and TT steps. The simulations also indicate that only the TA step can form a very efficient interbase hydrogen bond network in the TA-DNA conformation. Such an energetically stabilizing network is not achievable in the AA and TT steps. While it is viable in the AT step, structural constraints render the hydrogen bonding network energetically ineffective there.

DNA-Binding Proteins↗
PubMed1998

Progressive DNA bending is made possible by gradual changes in the torsion angle of the glycosyl bond.

Structural comparisons have led to the suggestion that the conformational rearrangement that would be required to change A-DNA into the TA-DNA form of DNA observed in the complex with the TATA box binding protein (TBP) could be completed by modifying only the value of the glycosyl bond chi by approximately 45 degrees. The lack of a high number of crystal structures of this type makes it difficult to conclude whether a smooth transition from A-DNA to TA-DNA can occur without disrupting at any point either the Watson-Crick base pairing or the A-DNA conformation of the backbone. To explore the possibility of such a smooth transition, constrained molecular dynamics simulations were carried out for the double-stranded dodecamer d(GGTATATAAAAC), in which a transition from A-DNA to TA-DNA was induced by modifying only the chi angle values. The results demonstrate the feasibility of a continuous path in the A-DNA to TA-DNA transition. Varying extents of DNA curvature are also attainable, by maintaining the A-DNA backbone structure and Watson-Crick hydrogen bonding while changing the chi angle value smoothly from that in A-DNA to one corresponding to B-DNA.

Base Composition↗
PubMed1997

The effect of the molecular mechanism of G protein-coupled receptor activation on the process of signal transduction.

A thermodynamic model of signal transduction that incorporates the possibility of multiple conformational states between the inactive and the active forms of the receptor was developed. The obtained equilibrium model is equivalent to the extended ternary complex of Samama et al. (J. Biol. Chem. 268 (1993) 4625-4636) if only two states of the receptor exist. These multiple equilibria between receptor states are modeled by two sets of equilibrium constants: K(piAR) and K(sigma piAR), in the presence of the ligand; and K(piR) and K(sigma piR), in the absence of the ligand. The higher the value of these constants, the more efficiently the active form of the receptor is generated. Intrinsic efficacy of the agonist is defined in the present formulation as the molecular processes induced by ligands in the receptor that lead to the active form of the receptor. Both the energetics (associated to K[piAR]) and mechanism of the process of receptor activation (associated to K[sigma piAR]) are important in eliciting the maximum response. Moreover, analytical expressions of basal activity, potency and maximum response were obtained. These definitions were used to classify the extra cellular ligand as agonists (K[sigma piAR] > K[sigma piR]), inverse agonists (K[sigma piR] > K[sigma piAR] > 0), neutral antagonists (K[sigma piAR] = K[sigma piR]), and pure antagonists.

Computer Simulation↗
PubMed1997

Does TATA matter? A structural exploration of the selectivity determinants in its complexes with TATA box-binding protein.

The binding of the TATA box-binding protein (TBP) to a TATA sequence in DNA is essential for eukaryotic basal transcription. TBP binds in the minor groove of DNA, causing a large distortion of the DNA helix. Given the apparent stereochemical equivalence of AT and TA basepairs in the minor groove, DNA deformability must play a significant role in binding site selection, because not all AT-rich sequences are bound effectively by TBP. To gain insight into the precise role that the properties of the TATA sequence have in determining the specificity of the DNA substrates of TBP, the solution structure and dynamics of seven DNA dodecamers have been studied by using molecular dynamics simulations. The analysis of the structural properties of basepair steps in these TATA sequences suggests a reason for the preference for alternating pyrimidine-purine (YR) sequences, but indicates that these properties cannot be the sole determinant of the sequence specificity of TBP. Rather, recognition depends on the interplay between the inherent deformability of the DNA and steric complementarity at the molecular interface.

Algorithms↗
PubMed1992

On the use of the transmembrane domain of bacteriorhodopsin as a template for modeling the three-dimensional structure of guanine nucleotide-binding regulatory protein-coupled receptors.

The molecular architecture of bacteriorhodopsin (BR) is commonly regarded as a structural template for the three-dimensional structure of membrane receptors that are functionally coupled to guanine nucleotide-binding regulatory proteins (GPCR). More recently, specific molecular models of such GPCR were constructed on the basis of the functional and structural relation of rhodopsin to BR as well as the sequence homology between rhodopsin and the GPCR. Such models of GPCR leave unresolved the difficulty caused by the apparent lack of any significant degree of sequence homology between the seven transmembrane helices (TMH) of BR and the portions in the sequence of the various GPCR that are considered to constitute their transmembrane domains. Evolutionary arguments offered in favor of the structural relation between BR and the opsins, and hence the GPCR, prompted our investigation of the possibility that the sequence homology, including any similarity in the distribution of kink-inducing proline residues among the helices, might have been obscured by the assumption that the TMH maintained their sequential order from BR in the evolution of the mammalian proteins. With a definition of the TMH in the neurotransmitter GPCR guided by hydropathicity predictions, and additional criteria used to define the span of each helix, optimal alignment of each pair of sequences was determined with no gaps allowed in the matching. The resulting alignment proposed here reveals considerable homology between the TMH in BR and those in GPCR, if the sequential order of the helices is ignored. These findings suggest the possibility that exon shuffling could have occurred in the proposed evolution of the GPCR gene from BR and point to a modification of the BR template to account for the correct packing of the helices in the tertiary structures of GPCR. These findings could guide the construction of three-dimensional models of the neurotransmitter GPCR on the basis of specific interhelical interactions observed in BR.

Amino Acid Sequence↗
PubMed1992

Theoretical studies on the histamine H2 receptor: molecular mechanism of action of antagonists.

The previously defined sites in the histamine H2 receptor model [Mol. Pharmacol. 40:980-987 (1991)] were used to elucidate the pharmacological mechanism of action of compounds that act as antagonists at the receptor. In this model, a formate anion is used both as the negative site at which the histamine cation is anchored to the receptor and as a proton acceptor site. An ammonium cation is used as a proton donor site. The proposed model of recognition of cimetidine, tiotidine, and ranitidine suggests that the monocationic form of the antagonists is the most favorable species to bind the receptor. Moreover, the mode of recognition follows the same trends obtained for compounds that act as agonists; the protonated site of the molecule, i.e., imidazolium in cimetidine, guanidinium in tiotidine, or substituted ammonium in ranitidine, anchors at the negative site of the receptor, whereas the nonbasic part, i.e., cyanoguanidine in cimetidine and tiotidine and nitrodiaminoethene in ranitidine, is located between the proton donor and acceptor sites. An energetic analysis of the interaction between the antagonists and the receptor model, including the energies of ligand desolvation, shows that histamine cannot compete effectively with cimetidine, tiotidine, or ranitidine for binding to the H2 receptor. The predicted order of antagonist potencies, based on differences of formation enthalpies (delta delta H1), reproduces qualitatively the experimental rank order.

Cimetidine↗
PubMed1991

Comparison of sucralfate and ranitidine in the treatment of duodenal ulcers.

Ninety patients with endoscopically proven duodenal ulcers, enrolled in this prospective, single-blind study, were randomized to sucralfate, 1 g q.i.d., 0.5 hour before meals on an empty stomach or at bedtime, or ranitidine, 150 mg b.i.d., for 4-8 weeks (phase I). Patients who healed during the treatment period were invited to participate in a maintenance therapy follow-up covering 1 year (phase II), during which they were treated with sucralfate, 1 g b.i.d. (before breakfast and at bedtime) or 150 mg of ranitidine at bedtime. After 4 weeks of treatment, healing rates were 30/40 (75.0%) with sucralfate and 36/42 (85.7%) with ranitidine, and healing rates were 39/40 (97.6%) and 40/42 (95.2%) respectively, after 8 weeks of treatment. At the end of the 6th and 12th months of phase II, respectively, relapse rates were 3/33 (9.4%) and 10/32 (31.3%) in the sucralfate group and 5/33 (15.2%) and 10/29 (34.5%) in the ranitidine group. Differences between sucralfate- and ranitidine-treated groups were not significant. Both treatments were well tolerated. We conclude that sucralfate is as effective and safe as ranitidine in the short-term treatment and prevention of relapse in patients with ulcer disease.

Administration, Oral↗
PubMed1991

Molecular mechanisms of radiation induced DNA damage: H-abstraction and beta-cleavage.

Quantum mechanical simulations of hydrogen abstraction by hydroxyl radical from methanol and ethanol yield barriers that agree very well with those measured experimentally. Analysis of the multiconfigurational wavefunction indicates that the strength of the C-H bond is the electronic parameter that has a major contribution to the barrier for H-abstraction. Similar analysis applied to 2-deoxy-D-ribose shows that the strength of a C-H bond together with the steric accessibility of the hydrogen determine that H4 is the most susceptible hydrogen for abstraction by a hydroxyl radical. Quantum mechanical simulations of beta-cleavage show that a concerted mechanism in which a water molecule assists in the bond breaking process is more likely than a SN1 mechanism. However, the polar transition state suggests that the environment of the DNA and the surrounding water will have an important effect on the reaction.

DNA↗
PubMed1991

Molecular mechanisms of radiation induced DNA damage: H-addition to bases, direct ionization and double strand break.

Structures and properties of G.(-H) and of TH. were obtained from quantum mechanical calculations. New AMBER parameters for these radicals were obtained to fit their structures and charge distributions. Molecular mechanics simulations of the conformational changes induced in a 12-mer of DNA, d(CGCGAATTCGCG), by these radicals show that the distances between the base and the C2' of the sugar becomes shorter. Such changes suggest that the base radical can abstract the H2' and transfer the radical from the base to the sugar. Once the radical becomes centered on the sugar a strand break can follow. A simultaneous formation of guanine and thymine radicals on opposite strands may lead to a double strand break.

Base Sequence↗
PubMed1991

Theoretical studies on the histamine H2 receptor: construction of a receptor model based on the structural properties of dimaprit and N alpha-guanylhistamine.

A histamine H2 receptor model was constructed based on the receptor sites previously proposed by Weinstein et al. [Mol. Pharmacol. 29:28-33 (1986)]. In this model, a glutamate or aspartate residue, simulated by a formate anion, is proposed both as the negative site at which the histamine cation is anchored to the receptor and as a proton-acceptor site. A proton-donor site, simulated by an ammonium cation, is proposed to model either a lysine, arginine, or histidine residue. The simulation of the activation mechanism of the histamine H2 receptor, inside the proposed receptor model, includes structure optimizations of stationary points and transition states with a split valence basis set. The proton movement from the proton donor site to the proton acceptor site, mediated by the imidazole ring of histamine, was found to be sequential in the potential energy surface. Results of the calculations reveal that both proton transfers are feasible from an energetical point of view. However, the proton movement from N(3) to the proton acceptor site has a higher energy of activation and, therefore, will be the rate-limiting step in the starting process that triggers the cascade of events that finally leads to a biological response. This model also provides a basis for explaining the molecular determinants of the pharmacological activity of N alpha-guanylhistamine. The structural properties of the guanidinium group allow N alpha-guanylhistamine to interact with the proposed receptor in two different modes. The proton-relay process, proposed as the trigger of the activation of the histamine H2 receptor, is likely to occur in only one of these binding modes. In the other case, N alpha-guanylhistamine acts as an antagonist because the barrier to proton transfer in this mode is too high. The partial agonism of N alpha-guanylhistamine is related to the ability of the drug to bind with the receptor in two different modes with similar affinity. An energetic analysis of the interaction between of ligand and the receptor model, including the energies of ligand desolvation, shows that histamine can compete with N alpha-guanylhistamine for the binding to the H2 receptor.

Computer Simulation↗
PubMed1989

Sucralfate versus ranitidine in the treatment of gastric ulcer. Randomized clinical results in short-term and maintenance therapy.

This clinical study compared the healing capacities of sucralfate and ranitidine in the treatment of gastric ulcer. Sixty patients were assigned at random to treatment with either sucralfate (1 g four times per day) or ranitidine (150 mg twice per day). The patients underwent endoscopy before inclusion in the study, after four weeks, and after eight weeks if the ulcers had not completely healed after the fourth week (phase I). Patients whose ulcers had healed were invited to participate in phase II, consisting of maintenance treatment for one year. The dosage was 1 g sucralfate twice per day or 150 mg ranitidine before going to bed. The patients underwent a clinical examination every three months and endoscopy every six months, and whenever symptoms suggested a relapse. After four weeks, the ulcers in 53 percent of the sucralfate-treated patients (16 of 30) had healed, compared with 56 percent of the ranitidine-treated patients (17 of 30). After eight weeks, the cumulative healing rates were 83 percent (25 of 30) and 86 percent (26 of 30), respectively. At the six-month follow-up visit, the relapse rates were seven of 21 (33.3 percent) in the sucralfate group and nine of 18 (50 percent) in the ranitidine group. After 12 months, the accumulative relapse rates were eight of 18 (44.4 percent) and nine of 18 (50.0 percent). The only side effects worth noting were mild constipation in four patients treated with sucralfate. One patient in the ranitidine group had myalgia and one reported headache in phase I. In conclusion, sucralfate appears to be as effective as ranitidine in the short-term treatment of gastric ulcers and in relapse prophylaxis.

Adult↗
PubMed1985

Dose effects of halothane on sensory evoked responses obtained from the cortex, reticular formation and central gray.

Sensory evoked field potentials were recorded from the mesencephalic reticular formation (MRF), central gray (CG) and somatosensory cortex (SCX), following incremental doses of halothane in freely-moving rats. Halothane concentrations of 0.25%, 0.5% 1.0% and 2.0% were used. In general, the responses from each structure were affected in dose response manner. The averaged acoustic evoked responses (AAER) exhibit more sensitivity to halothane than the averaged visual evoked responses (AVER). The evoked response and its components obtained from each structure were affected differently by halothane mainly following the initial two halothane doses, (0.25% and 0.5%); mainly increase in amplitude was observed in the recording obtained from the MRF, decrease in the CG, and mixed (increase and/or decrease) in SCX. The degree of the depression of the sensory evoked responses was directly correlated to the level of anesthesia as assessed by sural nerve stimulation.

Animals↗