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Biomedical subjects

P R Wills

Publications and source records attributed to P R Wills.

At least 19 recordsLinked to original sources
PubMed2001

Studies of solute self-association by sedimentation equilibrium: allowance for effects of thermodynamic non-ideality beyond the consequences of nearest-neighbor interactions.

A sedimentation equilibrium study of alpha-chymotrypsin self-association in acetate-chloride buffer, pH 4.1 I 0.05, has been used to illustrate determination of a dimerization constant under conditions where thermodynamic non-ideality is manifested beyond the consequences of nearest-neighbor interactions. Because the expressions for the experimentally determinable interaction parameters comprise a mixture of equilibrium constant and excluded volume terms, the assignment of reasonable magnitudes to the relevant virial coefficients describing non-associative cluster formation is essential for the evaluation of a reliable estimate of the dimerization constant. Determination of these excluded volume parameters by numerical integration over the potential-of-mean-force is shown to be preferable to their calculation by approximate analytical solutions of the integral for this relatively small enzyme monomer with high net charge (+10) under conditions of low ionic strength (0.05 M).

Algorithms↗
PubMed2000

Interpretation of thermodynamic non-ideality in sedimentation equilibrium experiments on proteins.

This investigation re-examines theoretical aspects of the allowance for effects of thermodynamic non-ideality on the sedimentation equilibrium distribution for a single macromolecular solute, and thereby resolves the question of the constraints that pertain to the definition of the activity coefficient term in the basic sedimentation equilibrium expression. Sedimentation equilibrium results for ovalbumin are then presented to illustrate a simple procedure for evaluating the net charge (valence) of a protein from the magnitude of the second virial coefficient in situations where the effective radius of the protein can be assigned. Finally, published sedimentation equilibrium results on lysozyme are reanalysed to demonstrate the feasibility of employing the dependence of the second virial coefficient upon ionic strength to evaluate both the valence and the effective radius of the non-interacting solute.

Animals↗
PubMed2000

Analysis of sedimentation equilibrium distributions reflecting nonideal macromolecular associations.

A rigorous statistical-mechanical approach is adopted to derive general quantitative expressions that allow for the effects of thermodynamic nonideality in equilibrium measurements reflecting interaction between dissimilar macromolecular reactants. An analytical procedure based on these expressions is then formulated for obtaining global estimates of equilibrium constants and the corresponding reference thermodynamic activities of the free reactants in each of several sedimentation equilibrium experiments. The method is demonstrated by application to results from an ultracentrifugal study of an electrostatic interaction between ovalbumin and cytochrome c (Winzor, D. J., M. P. Jacobsen, and P. R. Wills. 1998. Biochemistry. 37:2226-2233). It is demonstrated that reliable estimates of relevant thermodynamic parameters are extracted from the data through statistical analysis by means of a simple nonlinear fitting procedure.

Biophysical Phenomena↗
PubMed1998

Direct analysis of sedimentation equilibrium distributions reflecting complex formation between dissimilar reactants.

Procedures are developed for the characterization of thermodynamically ideal complex formation between dissimilar macromolecular reactants by direct analysis of sedimentation equilibrium distributions. Studies of an electrostatic interaction between ovalbumin and cytochrome c are used to illustrate the application of analyses pertaining to (i) the situation in which separate sedimentation equilibrium distributions for the two macromolecular constituents are available, (ii) that in which the experimental record reflects the distribution of only one constituent, and (iii) the situation in which a composite distribution for both constituents is the sole experimental record. An association constant of 63 000 (+/- 2000) M-1 is obtained for the 1:1 interaction between ovalbumin and cytochrome c under the conditions examined (pH 6.3, I 0.03). Because of their inherent simplicity, these direct analytical procedures offer potential for accommodating the effects of thermodynamic nonideality in dissimilar reactant systems.

Animals↗
PubMed1998

Selection dynamics in autocatalytic systems: templates replicating through binary ligation.

The theory of autocatalytic binary ligation is reviewed within the context of a consistently applied Michaelis-Menten quasi-steady-state approximation to obtain explicit analytical results describing time-course data from experiments. A detailed protocol for the step-wise elucidation of a minimal set of experimental parameters is outlined. The kinetic equations are then generalized to cases of self- and cross-catalysis among an arbitrary number of different templates and applied to experiments involving just two templates. Depending on the values of various kinetic parameters such systems can display exclusionary Darwinian selection corresponding to an exponential growth law, selective coexistence or coexistence of all species characteristic of a parabolic growth law; the intermediate behaviour arises as a property of the full mechanism analysed here. Our results are applicable to the classical case of self-replicating nucleic acids and their analogues as well as to newly discovered self-replicating peptides.

Catalysis↗
PubMed1997

The emergence of genetic coding in physical systems.

A simple model of molecular biological translation, based on the classification of polymers as either information carriers or functional catalysts, is used to analyse formal constraints on physical systems which utilise genetic coding. We investigate (i) how the structure-function relationship for coding assignment catalysts constrains the selection of genetic information which can sustain functional self-organisation and (ii) what general prerequisites must be satisfied for selection to give rise to an increase in functional complexity. This is done by considering two separate alphabets and defining the complete set of assignments from letters of one alphabet onto letters from the other. A code is defined as a set of assignments which maps each letter from the first alphabet onto a letter from the second alphabet. We enumerate all the embeddings of the assignment functions in the minimal sequence space of strings of letters from the second alphabet and demonstrate how the embeddings can be classified according to whether they allow different codes to be represented unambiguously in the minimal sequence space of strings of letters from the first alphabet. Non-minimal embeddings are also discussed. Finally, we consider how the mutual specification of letters of the two alphabets and assignment functions can be decomposed into more highly differentiated classes. Only a certain class of embeddings allows coding to be preserved under decomposition. We conclude that the evolution of increasing coding complexity can take place only when special conditions are satisfied regarding the structure-function relationship for the coding assignment catalysts.

Animals↗
PubMed1996

Thermodynamic analysis of the effects of small inert cosolutes in the ultracentrifugation of noninteracting proteins.

Considerations of the effect of a small cosolute on the sedimentation equilibrium distribution for a noninteracting protein have led to the development of a simple procedure for evaluating both the molecular weight of the protein and the second virial coefficient describing the excluded volume interaction between protein and cosolute. Its application is illustrated by analysis of sedimentation equilibrium distributions for bovine thyroglobulin and horse liver alcohol dehydrogenase in the presence of a range of sucrose concentrations, and also of those for aldolase in the presence of urea to obtain the subunit molecular weight of this tetrameric enzyme. The effects of sucrose concentration on the sedimentation coefficients of thyroglobulin, catalase, and horse liver alcohol dehydrogenase are also examined to demonstrate that the second virial coefficients for protein-cosolute excluded volume interaction may be determined, albeit with less precision, from the cosolute concentration required to render the sedimentation coefficient zero by virtue of its effect on the buoyancy term. These findings serve to reinforce the fact that the effects of small cosolutes usually ascribed to changes in "protein solvation" are envisaged more realistically in terms of excluded volume.

Alcohol Dehydrogenase↗
PubMed1995

Thermodynamic nonideality of enzyme solutions supplemented with inert solutes: yeast hexokinase revisited.

Published experimental results on the activating effect of polyethylene glycol on the interaction of yeast hexokinase with glucose (R.P. Rand, N.L. Fuller, P. Butko, G. Francis and P. Nicholls, Biochemistry, 32 (1993) 5925) are reinterpreted in statistical-mechanical terms of excluded volume. Of particular interest is the ability of this standard treatment of thermodynamic nonideality to accommodate the observed non-exponential dependence of the activation upon osmotic pressure of the polyethylene glycol solution--a dependence which is not predicted by analyses based on the concept of osmotic stress that was invoked originally to account for the results.

Enzyme Activation↗
PubMed1993

Self-organization of genetic coding.

The self-organization of genetic coding is studied in a simple model system which uses the products of translation as catalysts for the process. The system studied contains protein molecules chosen from a sequence space of high dimension. Catalysts which assign amino acids to codons are chosen in such a way that a random selection of proteins synthesizes further proteins randomly. Under some circumstances, dictated by the genetic information supplied to the system and the manner in which protein function depends on protein sequence, the state of random synthesis is unstable. The system then evolves spontaneously to a new state in which proteins synthesize further proteins in an ordered fashion, typically executing the rules of a simple code for the assignment of amino acids to codons. For some embeddings of protein functions in the protein sequence space, the domains of stability of the ordered and disordered states are calculated. Computer simulation verify that coding self-organization occurs in a variety of systems of the sort studied. Coding self-organization among catalysts which recognize genetic information is a high order co-operative selection process which provides the link between genotype and phenotype needed for the Darwinian evolution of complex biochemical systems.

Amino Acids↗
PubMed1993

Thermodynamic nonideality in macromolecular solutions: interpretation of virial coefficients.

Consideration is given to the interpretation of virial coefficients reflecting thermodynamic nonideality in incompressible solutions of a single macromolecular species for which there is no volume change on mixing. Expressions are presented for the concentration dependence of thermodynamic activity under conditions where either the chemical potential of solvent or the pressure is fixed, these two conditions being mutually exclusive. For the former situation, which applies to partition equilibrium procedures, the thermodynamic activity is most conveniently defined on the molar scale because the coefficients in polynomial expansions for the osmotic pressure and the activity coefficient in terms of molarity are then related to each other without the inclusion of partial molar volume terms. Under conditions of constant pressure a similar situation prevails provided that the osmotic pressure and the corresponding activity coefficient are expanded in powers of molality. In either case conversion of the virial expansions to the other concentration scale is possible, but requires the introduction of partial molar volume terms into the virial coefficients. The implications of these findings are discussed in relation to results obtained by osmometry, isopiestic measurements, equilibrium dialysis, gel chromatography, and sedimentation equilibrium.

Chromatography, Gel↗
PubMed1991

Transmissible familial Creutzfeldt-Jakob disease associated with five, seven, and eight extra octapeptide coding repeats in the PRNP gene.

The PRNP gene, encoding the amyloid precursor protein that is centrally involved in Creutzfeldt-Jakob disease (CJD), has an unstable region of five variant tandem octapeptide coding repeats between codons 51 and 91. We screened a total of 535 individuals for the presence of extra repeats in this region, including patients with sporadic and familial forms of spongiform encephalopathy, members of their families, other neurological and non-neurological patients, and normal controls. We identified three CJD families (in each of which the proband's disease was neuropathologically confirmed and experimentally transmitted to primates) that were heterozygous for alleles with 10, 12, or 13 repeats, some of which had "wobble" nucleotide substitutions. We also found one individual with 9 repeats and no nucleotide substitutions who had no evidence of neurological disease. These observations, together with data on published British patients with 11 and 14 repeats, strongly suggest that the occurrence of 10 or more octapeptide repeats in the encoded amyloid precursor protein predisposes to CJD.

Adult↗
PubMed1990

Stem loops in HIV and prion protein mRNAs.

Tat-dependent trans-activation in HIV requires presentation of a CUGGG pentanucleotide at the end of a stem loop within the tar site of the viral long terminal repeat. A tandem repeat within the open reading frame of the prion protein (PrP) mRNA is able to form similar stem loop structures with which the HIV tat protein could interact, disturbing PrP translation. Self-amplification of such a disturbance has been suggested as the cause of the scrapie group of diseases, including the scrapie-like human dementiae. The same mechanism may underly AIDS encephalopathy.

AIDS Dementia Complex↗
PubMed1989

The tetrameric form of ribosomal protein L7/L12 from Escherichia coli.

A tetrameric form of the ribosomal protein L7/L12 has been prepared and its structure studied by using hydrodynamic methods, photon correlation spectroscopy, and small angle x-ray scattering. The tetrameric nature of the protein preparation is confirmed by three independent determinations of its molecular weight, with analysis of accurate sedimentation equilibrium data giving the most reliable estimate. The species has a Stokes radius of 4.0 +/- 0.1 nm and an absolute frictional ratio of 1.7. Taken together, the hydrodynamic measurements suggest the possibility of a flat structure, and this is consistent with the x-ray scattering results. The molecule has a radius of gyration of 3.6 +/- 0.05 nm and a maximum dimension of 11-12 nm. A geometric model consisting of four elongated monomers, arranged in a plane, is proposed.

Bacterial Proteins↗
PubMed1989

Induced frameshifting mechanism of replication for an information-carrying scrapie prion.

A specific mistranslation mechanism for the replication of an infectious protein is described. The feedback mechanism requires the infectious agent to induce concerted frameshifts during the translation of a cellular gene. Each module of the tandem repeat region of the gene encoding the prion protein (PrP) associated with scrapie infectivity contains multiple sites of potential ribosomal frameshifting. It is proposed that some aberrant variants of PrP containing frameshifted peptides within the octapeptide repeat region of the protein backbone are able to replicate and cause scrapie by interfering with the translation and simultaneous translocation of nascent PrP molecules into the lumen of the endoplasmic reticulum. The model provides a plausible explanation for the behaviour of host-adapted scrapie strains as well as the aetiology of scrapie-like diseases. The hypothesis that a mistranslated PrP is the scrapie agent can also explain discrepancies between the published amino acid sequence of PrP and the sequence deduced from the gene.

Amino Acid Sequence↗