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Biomedical subjects

M Karplus

Publications and source records attributed to M Karplus.

At least 109 records · Page 6Linked to original sources
PubMed1996

Fundus pigmentation in retinopathy of prematurity.

OBJECTIVE: A prospective study design was used to investigate the association between different degrees of fundus pigmentation and the indicence of retinopathy of prematurity (ROP) among very low birth weight infants in a large neonatal intensive care unit. METHODS: The study group consisted of 161 infants weighing 1500 g or less at birth and included all infants born from 1988 to 1990 who survived at least 10 weeks. Presence or lack of any acute stage ROP was determined by weekly ophthalmological examination from the age of 4 weeks. The degree of fundus pigmentation was recorded for each infant during the first examination. RESULTS: Infants with dark fundus pigmentation were found to be at half the risk of developing ROP as compared with infants having light/medium fundus pigmentation (relative risk = 0.5; 95% confidence interval = 0.2-1.1). When controlled for birth weight, gestational age, length of oxygen therapy, and ethnic group in multivariate analysis, dark pigmentation was an independent and statistically significant protective factor (odds ration = 0.09, 95% confidence interval = 0.02-0.06). None of the infants with Stage III ROP or higher had dark pigmentation. CONCLUSION: We speculate that large amounts of melanin in the retinal pigment epithelium or choroid may protect the dark-pigmented very low birth weight infant from developing ROP.

Female↗
PubMed1995

The meaning of component analysis: decomposition of the free energy in terms of specific interactions.

Free energy simulations are of particular interest for the interpretation of macroscopic data in terms of microscopic interactions. This can be done by expressing calculated free energies as a sum of components that correspond to the contributions of different energy terms or different parts of the system. Since the resulting components depend on the integration path, care is required for their use. We show that a linear coupling scheme for the alchemical creation of a chemical identity corresponds to a particularly useful path because it leads to a symmetric decoupling of the free energy components. The path dependence also provides an additional degree of freedom that can be used to study different processes. This latter point is illustrated by a reinterpretation of a recent simulation on wild-type and mutant azurin by Mark and van Gunsteren.

Alchemy↗
PubMed1995

Acid and thermal denaturation of barnase investigated by molecular dynamics simulations.

The transition in barnase from the native state to a partially unfolded conformation has been studied by molecular dynamics simulations with explicit water molecules at 360 K and low pH(450 ps), and at 600 K and neutral pH (three simulations of 120, 250 and 200 ps each). The use of several simulations provides evidence that the results are not sensitive to initial conditions. To mimic low pH conditions, the acidic sidechains in barnase were neutralized and the two histidine residues were doubly protonated. Runs at 300 K showed that the solvated structures at low pH (300 ps) and neutral pH (310 ps) are very similar. The main structural differences involved the acidic residues, histidine residues, and the beta-turn connecting strands 4 and 5. When the temperature is raised to 360 K at low pH and to 600 K at neutral pH the barnase molecule begins to unfold. The molecule rapidly expands (Rg changes from 13.9 A to 15.3 A in 450 ps at 360 K and from 13.7 A to between 15.1 and 15.5 A in 120 ps at 600 K). However, the expansion is not uniform. In all the simulations, the chain termini, loops and the N-terminal parts of the main alpha-helix (helix 1) show a continuous and progressive unfolding. An essential step in the denaturation process is that the major alpha-helix (helix 1) separates from the beta-sheet; this is coupled to the exposure of the principal hydrophobic core, many of whose non-polar side chains become solvated by hydrogen-bonded water molecules. The barnase-water interaction energy improves during unfolding at the expense of the barnase self-energy. The deterioration of the intramolecular van der Waals energy suggests that the rupture of the tight packing during the initial unfolding phase contributes to the energy barrier of the denaturation process. The mutationally well-analyzed Asp8-Arg110-Asp12 double salt-bridge on the barnase surface is found to be marginally stable in the folded form in the simulations. A Poisson-Boltzmann calculation indicates that the salt-bridge is unstable; this is probably due to an overestimate of the solvation energy. A detailed analysis of the main hydrophobic core reveals that increase in solvent-accessible surface area and penetration of water molecules are simultaneous in the high-temperature simulation; at lower temperatures there is significant cavity formation and the entrance of the water molecules is somewhat delayed. The cavities occur in the neighborhood of the hydrophobic sidechains; the region formed by the sidechains of Val10, Leu14, Leu20, Tyr24, Ala74, Ile76 and Tyr90 is involved. The loosening of the core packing is coupled to an increase in the number of dihedral transitions.(ABSTRACT TRUNCATED AT 400 WORDS)

Bacterial Proteins↗
PubMed1995

Packaging of proteases and proteoglycans in the granules of mast cells and other hematopoietic cells. A cluster of histidines on mouse mast cell protease 7 regulates its binding to heparin serglycin proteoglycans.

Mouse mast cell protease 7 (mMCP-7) is a tryptase stored in the secretory granules of mast cells. At the granule pH of 5.5, mMCP-7 is fully active and is bound to heparin-containing serglycin proteoglycans. to understand the interaction of mMCP-7 with heparin inside and outside the mast cell, this trytase was first studied by comparative protein modeling. The "pro" form of mMCP-7 was then expressed in insect cells and studied by site-directed mutagenesis. Although mMCP-7 lacks known linear sequences of amino acis that interact with heparin, the three-dimensional model of mMCP-7 revealed an area on the surface of the folded protein away from the substrate-binding site that exhibits a strong positive electrostatic potential at the acidic pH of the granule. In agreement with this calculation, recombinant pro-mMCP-7 bound to a heparin-affinity column at pH 5.5 and readily dissociated from the column at pH > 6.5. Site-directed mutagenesis confirmed the prediction that the conversion of His residues 8,68, and 70 in the positively charged region into Glu prevents the binding of pro-mMCP-7 to heparin. Because the binding requires positively charged His residues, native mMCP-7 is able to dissociate from the protease/proteoglycan macromolecular complex when the complex is exocytosed from bone marrow-derived mast cells into a neutral pH environment. Many hematopoietic effector cells store positively charged proteins in granules that contain serglycin proteoglycans. The heparin/mMCP-7 interaction, which depends on the tertiary structure of the tryptase, may be representative of a general control mechanism by which hematopoietic cells maximize storage of properly folded, enzymatically active proteins in their granules.

Amino Acid Sequence↗
PubMed1995

pH dependence of binding reactions from free energy simulations and macroscopic continuum electrostatic calculations: application to 2'GMP/3'GMP binding to ribonuclease T1 and implications for catalysis.

An approach is described for extending free energy calculations to take into account the pH dependence of the relative binding of ligands to an enzyme or other receptor protein. The method is based on the calculation of the free energy difference for a single protonation state via the thermodynamic cycle simulation approach followed by inclusion of all possible protonation states of the enzyme and the inhibitor by use of a macroscopic continuum dielectric (Poisson-Boltzmann) model. A detailed formulation of the combined model is presented. It involves solution of the multiple equilibrium problem and makes use of the calculated pKa values of all titrating groups on both enzyme and ligand. The method is illustrated by calculations of the pH dependence of the differential binding of the inhibitors 2'GMP and 3'GMP to ribonuclease T1. A free energy simulation of the differential binding is made for a given protonation state of the enzyme and inhibitor. Although only qualitative agreement with experiment is obtained, the results provide insights concerning the interactions involved. The pH dependence of the binding is calculated by using the protonation state of the residues from the free energy simulation as the standard state for a Poisson-Boltzmann calculation. Information is obtained concerning the pKa values of the titrating amino acids in the free, 2'GMP and 3'GMP bound enzyme forms of RNase T1 and the difference in the pH dependence of the binding of 2'GMP and 3'GMP to RNase T1. The contributions of different types of interactions (e.g. protein residues versus solvent) to the free energy differences are examined. A free energy simulation of the pKa shift of Glu58 shows that it is important to consider both carboxyl oxygen atoms as possible protonation sites since they may behave very differently in a protein. It is found in the protein that the interactions with the solvent favor the neutral (protonated) state of Glu58. This contrasts sharply with the solution behavior, where the solvent favors the charged state. Analysis of the results shows that the interactions of bound water with other protein residues leads to the observed effect. Comparisons are made with a continuum calculation that uses the charged state employed in the free energy simulation.(ABSTRACT TRUNCATED AT 400 WORDS)

Catalysis↗
PubMed1995

Neural networks for secondary structure and structural class predictions.

A pair of neural network-based algorithms is presented for predicting the tertiary structural class and the secondary structure of proteins. Each algorithm realizes improvements in accuracy based on information provided by the other. Structural class prediction of proteins nonhomologous to any in the training set is improved significantly, from 62.3% to 73.9%, and secondary structure prediction accuracy improves slightly, from 62.26% to 62.64%. A number of aspects of neural network optimization and testing are examined. They include network overtraining and an output filter based on a rolling average. Secondary structure prediction results vary greatly depending on the particular proteins chosen for the training and test sets; consequently, an appropriate measure of accuracy reflects the more unbiased approach of "jackknife" cross-validation (testing each protein in the data-base individually).

Algorithms↗
PubMed1995

Zinc binding in proteins and solution: a simple but accurate nonbonded representation.

Force field parameters that use a combination of Lennard-Jones and electrostatic interactions are developed for divalent zinc and tested in solution and protein simulations. It is shown that the parameter set gives free energies of solution in good agreement with experiment. Molecular dynamics simulations of carboxypeptidase A and carbonic anhydrase are performed with these zinc parameters and the CHARMM 22 beta all-atom parameter set. The structural results are as accurate as those obtained in published simulations that use specifically bonded models for the zinc ion and the AMBER force field. The inclusion of longer-range electrostatic interactions by use of the Extended Electrostatics model is found to improve the equilibrium conformation of the active site It is concluded that the present parameter set, which permits different coordination geometries and ligand exchange for the zinc ion, can be employed effectively for both solution and protein simulations of zinc-containing systems.

Binding Sites↗
PubMed1995

Evaluation of comparative protein modeling by MODELLER.

We evaluate 3D models of human nucleoside diphosphate kinase, mouse cellular retinoic acid binding protein I, and human eosinophil neurotoxin that were calculated by MODELLER, a program for comparative protein modeling by satisfaction of spatial restraints. The models have good stereochemistry and are at least as similar to the crystallographic structures as the closest template structures. The largest errors occur in the regions that were not aligned correctly or where the template structures are not similar to the correct structure. These regions correspond predominantly to exposed loops, insertions of any length, and non-conserved side chains. When a template structure with more than 40% sequence identity to the target protein is available, the model is likely to have about 90% of the mainchain atoms modeled with an rms deviation from the X-ray structure of approximately 1 A, in large part because the templates are likely to be that similar to the X-ray structure of the target. This rms deviation is comparable to the overall differences between refined NMR and X-ray crystallography structures of the same protein.

Amino Acid Sequence↗
PubMed1995

An automated method for dynamic ligand design.

An automated method for the dynamic ligand design (DLD) for a binding site of known structure is described. The method can be used for the creation of de novo ligands and for the modification of existing ligands. The binding site is saturated with atoms (sp3 carbon atoms in the present implementation) that form molecules under the influence of a potential function that joins atoms to each other with the correct stereochemistry. The resulting molecules are linked to precomputed functional group minimum energy positions in the binding site. The generalized potential function allows atoms to sample a continuous parameter space that includes the Cartesian coordinates and their occupancy and type, e.g., the method allows change of an sp3 carbon into an sp2 carbon or oxygen. A parameter space formulated in this way can then be sampled and optimized by a variety of methods. In this work, molecules are generated by use of a Monte Carlo simulated annealing algorithm. The DLD method is illustrated by its application to the binding site of FK506 binding protein (FKBP), an immunophilin. De novo ligands are designed and modification of the immunosuppressant drug FK506 are suggested. The results demonstrate that the dynamic ligand design approach can automatically construct ligands which complement both the shape and charge distribution of the binding site.

Algorithms↗
PubMed1995

Breech presentation and cesarean section in term nulliparous women.

OBJECTIVE: To examine pregnancy outcome in nulliparous women with single term breech presentation. METHODS: Two departments of Obstetrics and Gynecology at the same hospital used different approaches to deliver nulliparous women with singleton breech presentation at term. One department (A) delivered by trial of labor and the other (B) delivered by elective cesarean section. Prospectively and blinded to obstetric condition, parturients were assigned to either department in a systematic alternate fashion. The study period covered 8 years (1985-1992). The pregnancy outcome parameters examined were: Apgar score, intra- and post-partum death and maternal and neonatal morbidity. Neonatal morbidity was classified in three major categories: non-neurological trauma, neurological signs and respiratory problems. RESULTS: The study included 264 women of whom 135 delivered in department A and 129 in department B. Department A had 35 vaginal and 100 cesarean births and department B 10 vaginal and 119 cesarean births. There was no intra-partum death and the only post-partum death occurred among vaginal deliveries. The Apgar score was significantly worse at 1 and 5 min in vaginally delivered babies of department B. Neonatal morbidity was significantly more frequent after vaginal births (P < 0.01). Maternal morbidity was significantly higher following cesarean sections (P < 0.01). Babies of vaginal deliveries had significantly higher non-neurological trauma (P < 0.01) and pathological neurological signs (P < 0.01) than those delivered by the abdominal route. CONCLUSION: The level of risk for mother and child in the nulliparous with term singleton breech, suggests cesarean section as the preferred route of delivery.

Adult↗
PubMed1995

Theoretical studies of protein folding and unfolding.

The mechanism of protein folding is being investigated theoretically by the use of both simplified and all-atom models of the polypeptide chain. Lattice heteropolymer simulations of the folding process have led to proposals for the folding mechanism and for the resolution of the Levinthal paradox. Both stability and rapid folding have been shown in model studies to result from the presence of a pronounced global energy minimum corresponding to the native state. Concomitantly, molecular dynamics simulations with detailed atomic models have been used to analyze the initial stages of protein unfolding. Results concerning possible folding intermediates and the role of water in the unfolding process have been obtained. The two types of theoretical approaches are providing information essential for an understanding of the mechanism of protein folding and are useful for the design of experiments to study the mechanism in different proteins.

Models, Theoretical↗
PubMed1995

Ion transport in the gramicidin channel: molecular dynamics study of single and double occupancy.

The structural and thermodynamic factors responsible for the singly and doubly occupied saturation states of the gramicidin channel are investigated with molecular dynamics simulations and free energy perturbation methods. The relative free energy of binding of all of the five common cations Li+, Na+, K+, Rb+, and Cs+ is calculated in the singly and doubly occupied channel and in bulk water. The atomic system, which includes the gramicidin channel, a model membrane made of neutral Lennard-Jones particles and 190 explicit water molecules to form the bulk region, is similar to the one used in previous work to calculate the free energy profile of a Na+ ion along the axis of the channel. In all of the calculations, the ions are positioned in the main binding sites located near the entrances of the channel. The calculations reveal that the doubly occupied state is relatively more favorable for the larger ions. Thermodynamic decomposition is used to show that the origin of the trend observed in the calculations is due to the loss of favorable interactions between the ion and the single file water molecules inside the channel. Small ions are better solvated by the internal water molecules in the singly occupied state than in the doubly occupied state; bigger ions are solvated almost as well in both occupation states. Water-channel interactions play a role in the channel response. The observed trends are related to general thermodynamical properties of electrolyte solutions.

Binding Sites↗
PubMed1995

Use of a minimum perturbation approach to predict TIM mutant structures.

A minimum perturbation conformational search approach is used to model the structures of the yeast triosephosphate isomerase (TIM) single mutant in which the catalytic base Glu165 is changed to Asp, and the double mutant in which Glu165 is changed to Asp and Ser96 to Pro. In chicken TIM this double mutant is referred to as a pseudo-revertant because some of the catalytic activity lost due to the first mutation is regained when the second mutation occurs. Three minimum energy structures were calculated for the Asp165 conformation in the yeast TIM single mutant and another three for the double mutant. One of the calculated minimum energy conformations for Asp165 in the E165D structure agrees well with the X-ray structure. However, this conformation is not that of the lowest energy and is not one of the three most common conformers for Asp found by Ponder and Richards. This suggests that when an amino acid is introduced it may not be able to conform to the more general rules that apply to protein structures of evolutionary origin. While the van der Waals energy largely determines the allowed minima, the relative ranking of the final minima is determined by electrostatic effects and can therefore be affected by the inclusion of crystal waters in the calculation. When the E165D calculation is repeated with an active-site water molecule fixed in its E165D X-ray structure position, the relative ranking of the minima shifts and the X-ray conformation for Asp165 is the lowest interaction energy conformer. Two of the E165D calculated minimum energy structures are essentially identical to two of the S96P/E165D minima. All of the calculated minima for both the E165D and S96P/E165D mutants position the Asp side chain such that the anti-orbital, and not the more basic syn-orbital, of the carboxylate would be utilized for proton abstraction. This observation may explain why the chicken TIM S96P/E165D mutant, for which the X-ray structure indicates that the syn-orbital is used, is a pseudo-revertant while the yeast TIM double mutant is not; no X-ray structure is available for the latter. The multiplicity of minima found in the present analysis makes clear that predicting the exact orientation of a single side chain is not as simple as might be expected.

Amino Acid Sequence↗
PubMed1994

Analysis of two-residue turns in proteins.

The conformational properties of tight two-residue beta-turns in proteins are examined by empirical energy function calculations. Twenty-five tight turns are studied in isolation, in the presence of the protein, and in the presence of the protein and crystal water molecules. The conformational properties are subdivided into those that are intrinsic to the turn and those that depend on the protein and water environment. Two factors are shown to determine the conformation of a tight beta-turn. One is the twist of the beta-sheet (responsible for selecting either a type I' or II' conformation as opposed to the more common types I or II) and the other is a local electrostatic effect (responsible for distinguishing between the type I' and II' conformations). In the rare cases where a two-residue turn is found in a type I conformation, there exists a stabilizing feature (turn-protein interaction, a side-chain in a conformation that stabilizes type I, etc.) which compensates for the unfavorable twist of the turn relative to the beta-sheet.

Amino Acids↗
PubMed1994

How does a protein fold?

The number of all possible conformations of a polypeptide chain is too large to be sampled exhaustively. Nevertheless, protein sequences do fold into unique native states in seconds (the Levinthal paradox). To determine how the Levinthal paradox is resolved, we use a lattice Monte Carlo model in which the global minimum (native state) is known. The necessary and sufficient condition for folding in this model is that the native state be a pronounced global minimum on the potential surface. This guarantees thermodynamic stability of the native state at a temperature where the chain does not get trapped in local minima. Folding starts by a rapid collapse from a random-coil state to a random semi-compact globule. It then proceeds by a slow, rate-determining search through the semi-compact states to find a transition state from which the chain folds rapidly to the native state. The elements of the folding mechanism that lead to the resolution of the Levinthal paradox are the reduced number of conformations that need to be searched in the semi-compact globule (approximately 10(10) versus approximately 10(16) for the random coil) and the existence of many (approximately 10(3)) transition states. The results have evolutionary implications and suggest principles for the folding of real proteins.

Models, Chemical↗
PubMed1994

The contribution of vibrational entropy to molecular association. The dimerization of insulin.

The association reaction of two molecules to form a single complex must overcome a large entropic barrier due to the loss of translational and rotational degrees of freedom; estimates of the T delta S term are of the order of 30 kcal/mol for proteins. The approach of Chandler and Pratt is used to provide a statistical mechanical formulation for the connection between the gas-phase and solution binding free energies. This makes possible a clear separation of the vibrational contribution to the gas-phase binding enthalpy and entropy from the solvation terms. Further, it suggests that the calculated gas-phase result, should be a good approximation in solution for many systems. To illustrate the formulation, a harmonic dynamics model is used to study the dimerization of insulin. The vibrational entropy increase in the dimer complex, relative to the two separate monomers, is 23 entropy units. This contributes -7.2 kcal/mol to the dimerization free energy. It is not possible to identify a small number of specific dimer modes that give rise to this entropy contribution. Instead, small alterations in the frequencies of many modes below 400 to 600 cm-1 are found to contribute. The relative importance of vibrational and other effects in macromolecule-macromolecule and macromolecule-small molecule associations is discussed.

Binding Sites↗
PubMed1994

Molecular recognition in proteins. Simulation analysis of substrate binding by a tyrosyl-tRNA synthetase mutant.

Alchemical molecular dynamics simulations are performed to determine the difference in the free energy of binding of the tyrosine substrate between the wild type of tyrosyl-tRNA synthetase (TyrRS) from Bacillus stearothermophilus and the mutant Tyr169-->Phe. The results are of general interest because the Tyr169 hydroxyl group interacts with the ammonium group of the substrate in a manner corresponding to that found in other amino acid binding proteins (e.g. the Asp receptor of the chemotactic bacterium Salmonella typhimurium and class I major histocompatibility complex molecules). The calculated free-energy change due to the Tyr169-->Phe mutation is 3.4 kcal/mol (the statistical error is +/- 0.5 kcal/mol) in satisfactory agreement with the experimental value of 3(+/- 0.5) kcal/mol. By use of thermodynamic integration, the contribution of the different terms to the free energy change are estimated. The path dependence of such a decomposition is discussed and it is suggested that the alchemical choice is of primary interest for understanding the interactions involved. There are large protein contributions to the alchemical free energy difference of the bound and free enzyme that cancel in the overall result. Due to this cancellation, the essential interactions contributing to the free-energy change are those between the OH group of Tyr169 and water in the free enzyme and those between the OH group of Tyr169 and the ammonium group of the substrate in the bound system. The results thus support simple models based on a balance of hydrogen bonding interactions.

Bacterial Proteins↗