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Biomedical subjects

H X Zhou

Publications and source records attributed to H X Zhou.

At least 19 recordsLinked to original sources
PubMed2001

The affinity-enhancing roles of flexible linkers in two-domain DNA-binding proteins.

Recently many attempts have been made to design high-affinity DNA-binding proteins by linking two domains. Here a theory for guiding these designs is presented. Flexible linkers may play three types of roles: (a) linking domains which by themselves are unfolded and bind to DNA only as a folded dimer (as in a designed single-chain Arc repressor), (b) connecting domains which can separately bind to DNA (as in the Oct-1 POU domain), and (c) linking a DNA-binding domain with a dimerization domain (as in the lambda repressor). In (a), the linker keeps the protein as a folded dimer so that it is always DNA-binding-competent. In (b), the linker is predicted to enhance DNA-binding affinity over those of the individual domains (with dissociation constants K(A) and K(B)) by p(d(0))/K(B) or p(d(0))/K(A), where p(d(0)) = (3/4pil(p)bL)(3/2) exp(-3d(0)(2)/4l(p)bL)(1 - 5l(p)/4bL +...) is the probability density for the end-to-end vector of the linker with L residues to have a distance d(0). In (c), the linker is predicted to enhance the binding affinity by K(d)(C)/p(d(0)), where K(d)(C) is the dimer dissociation constant for the dimerization domain. The predicted affinity enhancements are found to be actually reached by the Oct-1 POU domain and lambda repressor. However, there is room for improvement in many of the recently designed proteins. The theoretical limits presented should provide a useful guide for current efforts of designing DNA-binding proteins.

Binding Sites↗
PubMed2001

A unified picture of protein hydration: prediction of hydrodynamic properties from known structures.

Hydration is essential for the structural and functional integrity of globular proteins. How much hydration water is required for that integrity? A number of techniques such as X-ray diffraction, nuclear magnetic resonance (NMR) spectroscopy, calorimetry, infrared spectroscopy, and molecular dynamics (MD) simulations indicate that the hydration level is 0.3-0.5 g of water per gram of protein for medium sized proteins. Hydrodynamic properties, when accounted for by modeling proteins as ellipsoids, appear to give a wide range of hydration levels. In this paper we describe an alternative numerical technique for hydrodynamic calculations that takes account of the detailed protein structures. This is made possible by relating hydrodynamic properties (translational and rotational diffusion constants and intrinsic viscosity) to electrostatic properties (capacitance and polarizability). We show that the use of detailed protein structures in predicting hydrodynamic properties leads to hydration levels in agreement with other techniques. A unified picture of protein hydration emerges. There are preferred hydration sites around a protein surface. These sites are occupied nearly all the time, but by different water molecules at different times. Thus, though a given water molecule may have a very short residence time (approximately 100-500 ps from NMR spectroscopy and MD simulations) in a particular site, the site appears fully occupied in experiments in which time-averaged properties are measured.

Nuclear Magnetic Resonance, Biomolecular↗
PubMed2001

Stabilization of proteins in confined spaces.

We present theory showing that confining a protein to a small inert space (a "cage") should stabilize the protein against reversible unfolding. Examples of such spaces might include the pores within chromatography columns, the Anfinsen cage in chaperonins, the interiors of ribosomes, or regions of steric occlusion inside cells. Confinement eliminates some expanded configurations of the unfolded chain, shifting the equilibrium from the unfolded state toward the native state. The partition coefficient for a protein in a confined space is predicted to decrease significantly when the solvent is changed from native to denaturing conditions. Small cages are predicted to increase the stability of the native state by as much as 15 kcal/mol. Confinement may also increase the rates of protein or RNA folding.

Chaperonins↗
PubMed2001

Prediction of protein interaction sites from sequence profile and residue neighbor list.

Protein-protein interaction sites are predicted from a neural network with sequence profiles of neighboring residues and solvent exposure as input. The network was trained on 615 pairs of nonhomologous complex-forming proteins. Tested on a different set of 129 pairs of nonhomologous complex-forming proteins, 70% of the 11,004 predicted interface residues are actually located in the interfaces. These 7732 correctly predicted residues account for 65% of the 11,805 residues making up the 129 interfaces. The main strength of the network predictor lies in the fact that neighbor lists and solvent exposure are relatively insensitive to structural changes accompanying complex formation. As such, it performs equally well with bound or unbound structures of the proteins. For a set of 35 test proteins, when the input was calculated from the bound and unbound structures, the correct fractions of the predicted interface residues were 69 and 70%, respectively.

Computational Biology↗
PubMed2001

Fold recognition and accurate query-template alignment by a combination of PSI-BLAST and threading.

A homology-based structure prediction method ideally gives both a correct fold assignment and an accurate query-template alignment. In this article we show that the combination of two existing methods, PSI-BLAST and threading, leads to significant enhancement in the success rate of fold recognition. The combined approach, termed COBLATH, also yields much higher alignment accuracy than found in previous studies. It consists of two-way searches both by PSI-BLAST and by threading. In the PSI-BLAST portion, a query is used to search for hits in a library of potential templates and, conversely, each potential template is used to search for hits in a library of queries. In the threading portion, the scoring function is the sum of a sequence profile and a 6x6 substitution matrix between predicted query and known template secondary structure and solvent exposure. "Two-way" in threading means that the query's sequence profile is used to match the sequences of all potential templates and the sequence profiles of all potential templates are used to match the query's sequence. When tested on a set of 533 nonhomologous proteins, COBLATH was able to assign folds for 390 (73%). Among these 390 queries, 265 (68%) had root-mean-square deviations (RMSDs) of less than 8 A between predicted and actual structures. Such high success rate and accuracy make COBLATH an ideal tool for structural genomics.

Algorithms↗
PubMed2001

Disparate ionic-strength dependencies of on and off rates in protein-protein association.

Electrostatic interactions have been observed to play important roles in the kinetics of protein-protein association. Ionic strength, by its ability to modulate the magnitude of electrostatic interactions, has often been conveniently used to test their presence. From experiments on a wide range of associating proteins, a common feature has emerged: the on rates show strong dependence on ionic strength whereas the off rates are relatively insensitive. Here this feature is explained by an explicit description of a transition state for the association process and the suggestion that this transition is near the final bound state of two proteins. The molecular basis of the transition state in the bimolecular process lies in the fact that the bound state is characterized by local specific (e.g., van der Waals, hydrophobic, and electrostatic) interactions, whereas the unbound state is characterized by translational and rotational freedom. In the transition state the protein-protein pair encounters a free-energy maximum since its translational-rotational entropy is reduced while the specific interactions are not yet attained. In this formalism of the protein-protein association process, the enhancement of on rates by long-range electrostatic interactions can be written (analogous to an ordinary transition-state theory) in the form k(on) = k(0)(on)exp(-G(el)/k(B)T), where G(el) is the electrostatic free energy of the transition state.

Biopolymers↗
PubMed2001

Predicted structures of two proteins involved in human diseases.

Structures of 79 proteins involved in human diseases were predicted by sequence alignments with structural templates. The predicted structures for ALDP and CSA, proteins responsible for adrenoleukodystrophy and the Cockayne syndrome, respectively, were analyzed to elucidate the molecular basis of disease mutations. In particular we positioned residue P484 of ALDP in the homodimer interface. This positioning is consistent with a recent experimental finding that the mutation P484R significantly decreases the self-interaction of ALDP and suggests that the disease mechanism of this mutation lies in the impaired ALDP dimerization. We identified two new WD repeats in CSA and suggest that one of these forms part of the interaction surface with other proteins.

ATP Binding Cassette Transporter, Subfamily D, Mem↗
PubMed2000

Epidemiology of alveolar echinococcosis in Xinjiang Uygur autonomous region, China: a preliminary analysis.

The Xinjiang Uygur autonomous region (XUAR) in north-western China is known to be endemic for Echinococcosis multilocularis, the causative agent of human alveolar echinococcosis (HAE). Despite regular reports of HAE cases in their region, very little is known about the local epidemiology of the disease or the transmission of E. multilocularis. The epidemiology of HAE in Xinjiang has now been investigated by the analysis of data collected from the medical records of 157 clinical cases who had attended the four main hospitals in the region. These data indicate that the disease is relatively common in the Altai, western Junggar, and Tianshan mountain ranges, whereas the Tarim and Junggar basins are likely to be of low endemicity. The prevalence of the disease in the Kunlun mountains is not clear. Semi-nomadic groups, especially those of Kazakh or Mongol origin, have a higher risk of infection than other ethnic groups. Prevalence of the disease in humans appears to be correlated with aspects of the local climate, such as annual precipitation and temperature. Red foxes, Microtinae, Ochotona spp. and Marmota spp. may be important hosts of E. multilocularis in the XUAR, sustaining the transmission cycles of the parasite.

Adolescent↗
PubMed1999

Hydrogen bonds between short polar side chains and peptide backbone: prevalence in proteins and effects on helix-forming propensities.

A survey of 322 proteins showed that the short polar (SP) side chains of four residues, Thr, Ser, Asp, and Asn, have a very strong tendency to form hydrogen bonds with neighboring backbone amides. Specifically, 32% of Thr, 29% of Ser, 26% of Asp, and 19% of Asn engage in such hydrogen bonds. When an SP residue caps the N terminal of a helix, the contribution to helix stability by a hydrogen bond with the amide of the N3 or N2 residue is well established. When an SP residue is in the middle of a helix, the side chain is unlikely to form hydrogen bonds with neighboring backbone amides for steric and geometric reasons. In essence the SP side chain competes with the backbone carbonyl for the same hydrogen-bonding partner (i.e., the backbone amide) and thus SP residues tend to break backbone carbonyl-amide hydrogen bonds. The proposition that this is the origin for the low propensities of SP residues in the middle of alpha helices (relative to those of nonpolar residues) was tested. The combined effects of restricting side-chain rotamer conformations (documented by Creamer and Rose, Proc Acad Sci USA, 1992;89:5937-5941; Proteins, 1994;19:85-97) and excluding side- chain to backbone hydrogen bonds by the helix were quantitatively analyzed. These were found to correlate strongly with four experimentally determined scales of helix-forming propensities. The correlation coefficients ranged from 0.72 to 0.87, which are comparable to those found for nonpolar residues (for which only the loss of side-chain conformational entropy needs to be considered).

Asparagine↗
PubMed1998

Conformation gating as a mechanism for enzyme specificity.

Acetylcholinesterase, with an active site located at the bottom of a narrow and deep gorge, provides a striking example of enzymes with buried active sites. Recent molecular dynamics simulations showed that reorientation of five aromatic rings leads to rapid opening and closing of the gate to the active site. In the present study the molecular dynamics trajectory is used to quantitatively analyze the effect of the gate on the substrate binding rate constant. For a 2. 4-A probe modeling acetylcholine, the gate is open only 2.4% of the time, but the quantitative analysis reveals that the substrate binding rate is slowed by merely a factor of 2. We rationalize this result by noting that the substrate, by virtue of Brownian motion, will make repeated attempts to enter the gate each time it is near the gate. If the gate is rapidly switching between the open and closed states, one of these attempts will coincide with an open state, and then the substrate succeeds in entering the gate. However, there is a limit on the extent to which rapid gating dynamics can compensate for the small equilibrium probability of the open state. Thus the gate is effective in reducing the binding rate for a ligand 0.4 A bulkier by three orders of magnitude. This relationship suggests a mechanism for achieving enzyme specificity without sacrificing efficiency.

Acetylcholine↗
PubMed1998

Electrostatic enhancement of diffusion-controlled protein-protein association: comparison of theory and experiment on barnase and barstar.

The electrostatic enhancement of the association rate of barnase and barstar is calculated using a transition-state theory like expression and atomic-detail modeling of the protein molecules. This expression predicts that the rate enhancement is simply the average Boltzmann factor in the region of configurational space where association occurs instantaneously in the diffusion-controlled limit. Based on experimental evidence, this "transition state" is defined by configurations in which, relative to the stereospecifically bound complex, the two proteins are shifted apart by approximately 8 A (so a layer of water can be accommodated in the interface) and the two binding surfaces are rotated away by 0 degrees to 3 degrees. The values of the average Boltzmann factor, calculated by solving the Poisson-Boltzmann equation, for the wild-type complex and 16 complexes with single mutations are found to correlate well with experimental results for the electrostatic rate enhancement. The predicted rate enhancement is found to be somewhat insensitive to the precise definition of the transition state, due to the long-range nature of electrostatic interactions. The experimental ionic strength dependence of the rate enhancement is also reasonably reproduced.

Bacterial Proteins↗
PubMed1998

Correlation between rate of enzyme-substrate diffusional encounter and average Boltzmann factor around active site.

The utility of the average Boltzmann factor around the active site of an enzyme as the predictor of the electrostatic enhancement of the substrate binding rate is tested on a set of data on wild-type acetylcholinesterase and 18 charge mutants recently obtained by Brownian dynamics simulations. A good correlation between the average Boltzmann factors and the substrate binding rate constants is found. The effects of single charge mutations on both the Boltzmann factor and the substrate binding rate constant are modest, i.e., < 5 fold increase or decrease. This is consistent with the experimental results of Shafferman et al. but does not support their suggestion that the overall rate of the catalytic reaction is not limited by the diffusional encounter of acetylcholinesterase and its substrate.

Acetylcholine↗
PubMed1997

Design of fast enzymes by optimizing interaction potential in active site.

The diffusional encounter between substrate and enzyme, and hence catalytic efficiency, can be enhanced by mutating charged residues on the surface of the enzyme. In this paper we present a simple method for screening such mutations. This is based on our earlier result that electrostatic enhancement of the enzyme-substrate binding rate constant can be accounted for just by the interaction potential within the active site. Assuming that catalytic and structural integrity is maintained, the catalytic efficiency can be optimized by surface charge mutations which lead to stronger interaction potential within the active site. Application of the screening method on superoxide dismutase shows that only charge mutations close to the active site will have practical effect on the catalytic efficiency. This rationalizes a large number of findings obtained in previous simulation and experimental studies.

Animals↗
PubMed1997

Modeling of protein conformational fluctuations in pKa predictions.

A method is presented to account for conformational fluctuations of a protein in predicting the pK(a) values of its titrating groups. Conformations of the protein are generated by conventional molecular dynamics or Monte Carlo simulations, in which the protonations of the titrating groups are fixed. For each protein conformation, the electrostatic free energies required to add a proton charge to a titrating group while other groups are either unprotonated or protonated are calculated within a dielectric continuum model. These are used to determine the mean protonations of the titrating groups in the conformation at a series of pH values. The mean protonations are then used to determine the relative weight of the particular conformation with the titrating groups having all possible protonations. A conformationally averaged mean protonation for each titrating group is finally obtained by the weighted sum of the group's mean protonations in all the conformations. This method is applied to yeast iso-1-ferricytochrome c. The predicted pK(a) values are in general agreement with experimental results.

Computer Simulation↗
PubMed1997

Enhancement of protein-protein association rate by interaction potential: accuracy of prediction based on local Boltzmann factor.

Electrostatic interactions are known experimentally to enhance the rate of protein-protein association by three to four orders of magnitude. However, theoretical efforts to quantitatively account for such rate enhancement have been hampered by the need to consider a large number of relative configurations of two associating proteins sampled during their diffusional encounter. Our recent work indicates that a good estimate of the rate enhancement is given by the average Boltzmann factor in the region of configurational space where association can effectively take place. This estimate is tested on a model system consisting of two spherical proteins, each with a "reactive patch." Three different forms of interaction potential are considered. Comparison with exact results for the association rate constant demonstrates that predictions based on the local Boltzmann factor are accurate to within approximately 50% for realistic sizes of the reactive region and amplitudes of the interaction potential.

Computer Simulation↗
PubMed1996

Theory and simulation of the time-dependent rate coefficients of diffusion-influenced reactions.

A general formalism is developed for calculating the time-dependent rate coefficient k(t) of an irreversible diffusion-influenced reaction. This formalism allows one to treat most factors that affect k(t), including rotational Brownian motion and conformational gating of reactant molecules and orientation constraint for product formation. At long times k(t) is shown to have the asymptotic expansion k(infinity)[1 + k(infinity) (pie Dt)-1/2 /4 pie D + ...], where D is the relative translational diffusion constant. An approximate analytical method for calculating k(t) is presented. This is based on the approximation that the probability density of the reactant pair in the reactive region keeps the equilibrium distribution but with a decreasing amplitude. The rate coefficient then is determined by the Green function in the absence of chemical reaction. Within the framework of this approximation, two general relations are obtained. The first relation allows the rate coefficient for an arbitrary amplitude of the reactivity to be found if the rate coefficient for one amplitude of the reactivity is known. The second relation allows the rate coefficient in the presence of conformational gating to be found from that in the absence of conformational gating. The ratio k(t)/k(0) is shown to be the survival probability of the reactant pair at time t starting from an initial distribution that is localized in the reactive region. This relation forms the basis of the calculation of k(t) through Brownian dynamics simulations. Two simulation procedures involving the propagation of nonreactive trajectories initiated only from the reactive region are described and illustrated on a model system. Both analytical and simulation results demonstrate the accuracy of the equilibrium-distribution approximation method.

Algorithms↗