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D Bashford

Publications and source records attributed to D Bashford.

7 recordsLinked to original sources
PubMed1992

Electrostatic calculations of the pKa values of ionizable groups in bacteriorhodopsin.

The effects of solvation and charge-charge interactions on the pKa of ionizable groups in bacteriorhodopsin have been studied using a macroscopic dielectric model with atom-level detail. The calculations are based on the atomic model for bacteriorhodopsin recently proposed by Henderson et al. Even if the structural data are not resolved at the atomic level, such calculations can indicate the quality of the model, outline some general aspects of electrostatic interactions in membrane proteins, and predict some features. The effects of structural uncertainties on the calculations have been investigated by conformational sampling. The results are in reasonable agreement with experimental measurements of several unusually large pKa shifts (e.g. the experimental findings that Asp96 and Asp115 are protonated in the ground state over a wide pH range). In general, we find that the large unfavorable desolvation energies of forming charges in the protein interior must be compensated by strong favorable charge-charge interactions, with the result that the titrations of many ionizable groups are strongly coupled to each other. We find several instances of complex titration behavior due to strong electrostatic interactions between titrating sites, and suggest that such behavior may be common in proton transfer systems. We also propose that they can help to resolve structural ambiguities in the currently available density map. In particular, we find better agreement between theory and experiment when a structural ambiguity in the position of the Arg82 side-chain is resolved in favor of a position near the Schiff base.

Bacteriorhodopsins↗
PubMed1990

pKa's of ionizable groups in proteins: atomic detail from a continuum electrostatic model.

A macroscopic electrostatic model is used to calculate the pKa values of the titratable groups in lysozyme. The model makes use of detailed structural information and treats solvation self-energies and interactions arising from permanent partial charges and titratable charges. Both the tetragonal and triclinic crystal structures are analyzed. Half of the experimentally observed pKa shifts (11 out of 21) are well reproduced by calculations for both structures; this includes the unusually high pKa of Glu 35 in the active site. For more than half the pKa's (13 out of 21), there is a large difference (1-3.3 pK units) between the results from the two structures. Many of these correspond to the titrating groups for which the calculations are in error. Since for an ionic strength of 0.1 M the Debye screening between titratable groups leads to a very high effective dielectric constant (the average value for all pairs of titrating groups is approximately 900), near-neighbor interactions dominate the pKa perturbations. Thus, the pKa values are very sensitive to the details of the local protein conformation, and it is likely that side-chain mobility has an important role in determining the observed pKa shifts.

Computer Simulation↗
PubMed1988

Electrostatic effects of charge perturbations introduced by metal oxidation in proteins. A theoretical analysis.

A macroscopic dielectric model for the interactions between charges in proteins is used to calculate the changes in His residue pKa values induced in azurin by oxidation of the copper. The calculated results agree with nuclear magnetic resonance experiments to within the uncertainty associated with the measurements. It is found that a large apparent dielectric constant can describe the interaction between two protein groups, even if the shortest path between them is through the protein, which is assumed to have a low dielectric constant.

Azurin↗
PubMed1988

Diffusion-collision model for the folding kinetics of myoglobin.

The diffusion-collision model has been used to analyze the folding kinetics of myoglobin. The microdomains, which are the basic units that coalesce during the folding, are identified with the helices and the stabilizing contacts between helices are determined from the native structure. Both association and dissociation reactions are included and a range of stabilization parameters is investigated to determine the variation in overall rate and the relative contributions made by different intermediates during the folding process. In a comparison of folding to the native state and to the midpoint of the folding transition (i.e., 50% native protein at the completion of the reaction) significant differences in the contributing intermediates are found.

Diffusion↗
PubMed1987

Determinants of a protein fold. Unique features of the globin amino acid sequences.

The three-dimensional structures of globins are known, from crystallographic analyses, to be very similar. Their amino acid sequences, however, differ greatly. Only two residues are absolutely conserved in all sequences, and the residue identities of some pairs of sequences are only 16%. We have determined the nature and exact extent of the sequence variations and the extent to which the conserved features of the globin sequences are unique to this family. The 226 globin sequences now known were aligned and analysed. Because distantly related protein sequences cannot be aligned correctly without the use of structural data, we developed a method that incorporated structural information into the alignment procedure. Analysis of the aligned sequences show that: (1) Although individual chains vary in size between 132 and 157 residues, deletions and insertions result in there being only 102 residue sites common to all globins. These sites form six separate regions. Insertions and deletions between these regions means that their separations can vary in different sequences. (2) Within the conserved regions there are 32 sites that almost always contain hydrophobic residues. In the known structures, these sites are in the protein interior. We measured the variations in the size of the residues that occur in the 226 sequences at these sites. At six sites the residues differ in size by less than 40 A3, at 11 sites they differ by 40 to 100 A3, and at 15 sites they differ by more than 100 A3. There are two other conserved buried sites: one contains the His linked to the haem iron and the other usually contains a His involved with the haem ligand. (3) Within the conserved regions there are another 32 sites that are almost always occupied by charged, polar or small non-polar (Gly or Ala) residues. In the known structures, these sites are on the protein surface. To determine the extent to which the conserved features found for the globin sequences are unique to that protein family, the following procedure was used. The six conserved regions, and the residue restrictions that occur at the 66 sites within these regions, were encoded into two "templates". One was based only on the sequences so far determined; the other was extended to include as yet unobserved substitutions that seemed plausible on the basis of size, hydrophobicity and polarity. Each of the 3286 non-globin sequences in the data bank was then examined by a computer program to see how closely it could be matched to these templates.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1984

Diffusion-collision model for the folding kinetics of the lambda-repressor operator-binding domain.

The operator-binding domain of the lambda-repressor contains five alpha-helices and an extended N-terminal arm in the crystal structure determined by Pabo and Lewis reported in Nature 298, 443, 1982 (1). The four helices form a "box" enclosing a hydrophobic core with the fifth helix interacting with the equivalent helix in a dimer. With a small number of well-defined secondary structure elements (microdomains), the repressor is well suited for an analysis of its folding pathways and kinetics by use of the diffusion-collision model. In this paper, the basic elements of the model appropriate to a several microdomain protein are formulated and applied to a set of folding pathways consistent with the crystal structure of the operator-binding domain. The overall kinetics, as well as the time-dependence of intermediate states are determined as a function of the microdomain stability parameter.

Binding Sites↗