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P D Ross

Publications and source records attributed to P D Ross.

At least 19 recordsLinked to original sources
PubMed1992

Conformational changes of a viral capsid protein. Thermodynamic rationale for proteolytic regulation of bacteriophage T4 capsid expansion, co-operativity, and super-stabilization by soc binding.

We have used differential scanning calorimetry in conjunction with cryo-electron microscopy to investigate the conformational transitions undergone by the maturing capsid of phage T4. Its precursor shell is composed primarily of gp23 (521 residues): cleavage of gp23 to gp23* (residues 66 to 521) facilitates a concerted conformational change in which the particle expands substantially, and is greatly stabilized. We have now characterized the intermediate states of capsid maturation; namely, the cleaved/unexpanded, state, which denatures at tm = 60 degrees C, and the uncleaved/expanded state, for which tm = 70 degrees C. When compared with the precursor uncleaved/unexpanded state (tm = 65 degrees C), and the mature cleaved/expanded state (tm = 83 degrees C, if complete cleavage precedes expansion), it follows that expansion of the cleaved precursor (delta tm approximately +23 degrees C) is the major stabilizing event in capsid maturation. These observations also suggest an advantage conferred by capsid protein cleavage (some other phage capsids expand without cleavage): if the gp23-delta domains (residues 1 to 65) are not removed by proteolysis, they impede formation of the stablest possible bonding arrangement when expansion occurs, most likely by becoming trapped at the interface between neighboring subunits or capsomers. Icosahedral capsids denature at essentially the same temperatures as tubular polymorphic variants (polyheads) for the same state of the surface lattice. However, the thermal transitions of capsids are considerably sharper, i.e. more co-operative, than those of polyheads, which we attribute to capsids being closed, not open-ended. In both cases, binding of the accessory protein soc around the threefold sites on the outer surface of the expanded surface lattice results in a substantial further stabilization (delta tm = +5 degrees C). The interfaces between capsomers appear to be relatively weak points that are reinforced by clamp-like binding of soc. These results imply that the "triplex" proteins of other viruses (their structural counterparts of soc) are likely also to be involved in capsid stabilization. Cryo-electron microscopy was used to make conclusive interpretations of endotherms in terms of denaturation events. These data also revealed that the cleaved/unexpanded capsid has an angular polyhedral morphology and has a pronounced relief on its outer surface. Moreover, it is 14% smaller in linear dimensions than the cleaved/expanded capsid, and its shell is commensurately thicker.

Allosteric Regulation↗
PubMed1992

Thermodynamics of Cro protein-DNA interactions.

Using a highly sensitive pulsed-flow microcalorimeter, we have measured the changes in enthalpy and determined the thermodynamic parameters delta H, delta S degree, delta G degree, and delta C(p) for Cro protein-DNA association reactions. The reactions studied include sequence-nonspecific DNA association and sequence-specific DNA associations involving single- and multiple-base alterations and/or single-amino acid alteration mutants. (i) The association of Cro protein with nonspecific DNA at 15 degrees C is characterized by delta H = +4.4 kcal.mol-1 (1 cal = 4.18J), delta S degrees = 49 cal.mol-1.K-1, delta G degrees = -9.7 kcal.mol-1, and delta Cp congruent to 0; the association with specific high-affinity operator OR3 DNA is characterized by delta H = +0.8 kcal.mol-1, delta S degree = 59 cal.mol-1.K-1, delta G degree = -16.1 kcal.mol-1, and delta Cp = -360 cal.mol-1.K-1, respectively. Both nonspecific and specific Cro-DNA associations are entropy-driven. (ii) Plots of delta H vs. delta Cp and delta S degree vs. delta Cp for the 20 association reactions studied fall into two correlation groups with linear slopes of +9.4 K and -20.5 K and of -0.03 and -0.14, respectively. These regression lines have common intercepts, at the delta H and delta S degree values of nonspecific association (where delta Cp congruent to 0). The results suggest that there are, at least, two distinct conformational subclasses in specific Cro-DNA complexes, stabilized by different combinations of enthalpic and entropic contributions. The delta G degree and delta Cp values form an approximately single linear correlation group as a consequence of compensatory contributions from delta H and delta S degree to delta G degree and to delta Cp. Cro protein-DNA associations share some similar thermodynamic properties with protein folding, but their overall energetics are quite different. Although the nonspecific complex is stabilized predominantly by electrostatic forces, it appears that H bonds, van der Waals contacts, hydrophobic effects, and charge interactions all contribute to the stability (delta G degree and delta Cp) of the specific complex. (iii) The variations in the values of the thermodynamic parameters are in general accord with our knowledge of the structure of the Cro-DNA complex.

Base Sequence↗
PubMed1992

Origins and consequences of ligand-induced multiphasic thermal protein denaturation.

The presence of subsaturating levels of a high-affinity ligand has been demonstrated both by experiment and calculation to have far-reaching consequences on thermally induced protein denaturation due to the coupling between the protein denaturation and ligand-binding equilibria. Under such circumstances, a protein may undergo biphasic denaturation even though in the absence of ligand it exhibits a thermogram comprised of a single essentially symmetric endotherm. Up to now, the presence of just 2 maxima in the thermogram has been presented merely as an experimental observation or as the result of equilibrium computations. Here we develop a thermodynamic description of the linkage between these equilibria in which the number of cusps present in the thermogram correlates with the number of resolved steps in the plot of saturation level of remaining native protein vs temperature (i.e., the thermal binding curve). During thermally induced denaturation, the concentration of native protein decreases; thus, the native protein in effect is titrated with ligand at constant total ligand concentration. The free ligand concentration The free ligand concentration increases substantially through the release of bound ligand by unfolding protein thereby increasing the saturation level of the remaining native protein. The form of this thermal binding curve is a function of the number of ligand-binding sites on the protein, the magnitudes of the association constants, and the total ligand and total protein concentrations. As a result, the model predicts multiphasic denaturation of a single cooperative unit when the thermal binding curve consists of discrete multiple steps. The presence of only 2 maxima (i.e., a single cusp) in a thermogram for a protein with multiple sites on the native species derives from the form of the thermal binding curve, which in this case is a single-step sigmoidal plot, and not from the predominant denaturation of unliganded and fully liganded native species. In addition, it is shown that, in general, the contributions from the denaturation of individual native protein species are decidedly non-two-state in character; thus, simple deconvolution should not be carried out. The effects of nonzero values of delta Cp and d delta Cp/dT for denaturation and of changes in enthalpy and in heat capacity for ligand binding, as well as the interaction of ligand with the denatured protein, are explored also.

Binding Sites↗
PubMed1992

Relationship between bone mass and rates of bone change at appendicular measurement sites.

The rate of bone change among postmenopausal women may vary depending upon the initial bone mass. Examining this possibility is difficult, however, because of a negative statistical bias that occurs when change is regressed against the initial value of the same variable. In this article, four statistical methods were applied to measure the association between bone mass and the rate of bone change. The study population was Japanese-American women, who were monitored for approximately 5 years. Bone changes were determined for the calcaneus and the distal and proximal radius. The results were consistent across the bone sites but differed between statistical methods. Three of the four methods indicated that the women with the greater bone mass had the greater loss rates. The fourth method did not support this association. Possible reasons for the discordant results are discussed. Using the "best" estimate of the relationship, a gradual convergence of bone mass was projected over time toward the population mean. The convergence occurred because women with higher bone mass had a somewhat faster loss rate than women with lower bone mass. Overall, however, the variation in bone mass between individuals was large compared to the rate of convergence.

Absorptiometry, Photon↗
PubMed1992

Ability of vertebral dimensions from a single radiograph to identify fractures.

It has been proposed that vertebral dimensions be used to objectively identify vertebral fractures, permitting standardization of methodology for comparisons between studies. In this report, we evaluate the ability of various vertebral dimensions and ratios to identify "abnormal" vertebrae. As no "gold standard" exists for prevalent vertebral fractures, we examined the ability of cross-sectional dimensions (at a single point in time) to detect fractured vertebrae that had been identified from changes in dimensions compared with previous radiographs. Theoretically, a cutoff of 3 SD below the mean will rarely misclassify normal vertebrae as fractured (specificity = 99.9%). However, we found that this cutoff correctly identified only about 70% of the incident fractures. A less stringent criterion (2 SD below the mean; theoretical specificity = 97.7%) identified about 85-90% of true fractures. Dividing by stature or other vertebral heights sometimes yielded marginal improvements in the ability of the anterior or posterior height dimensions to diagnose fractures. The results suggest that the true fracture prevalence may sometimes be substantially higher than suggested by cross-sectional vertebral measurements.

Bone and Bones↗
PubMed1992

A method for estimating the uncertainty of future bone mass.

The development of statistical models for estimating fracture probability is a promising method for quantitating and optimizing the clinical utility of bone mass measurements. Earlier models have assumed that future bone mass could be predicted exactly and were, therefore, limited to analyses that assume the loss rate is known in advance. Since bone loss rates may vary over time and cannot be predicted accurately, we have developed a new model, based on empirical data, that estimates the degree of uncertainty associated with predicted bone mass. Without a bone mass measurement, the population mean must be assumed for an individual. For the calcaneus, the standard deviation of the population distribution is about 60 mg/cm2. By measuring bone mass, one can determine how close or far from the mean an individual's true bone mass is, with a standard deviation (SD) of about 3 mg/cm2. Without a subsequent bone mass measurement, our model predicts that the uncertainty (standard deviation) in calcaneal bone mass will increase approximately sixfold (relative to the reproducibility at the initial measurement) over a period of five years for women under age 60, from 3 mg/cm2 to 19 mg/cm2. The five-year increase in uncertainty is approximately fourfold for women over age 60, from 3 to 13 mg/cm2. However, the uncertainty in bone mass for an individual five years after the initial measurement is still only one third to one fifth that of the entire population, and can be reduced to the initial level by obtaining another measurement. Furthermore, the predicted (or measured) values are usually much better estimates of an individual's true bone mass than simply assuming the population average.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1991

Thermodynamics of antiparallel hairpin-double helix equilibria in DNA oligonucleotides from equilibrium ultracentrifugation.

Five highly palindromic DNA dodecamers, four of which may form G-A or I-A purine-purine mispairs at either the 5.8 or 6.7 positions, have been studied at sedimentation equilibrium in the analytical ultracentrifuge. Each DNA oligonucleotide forms an equilibrium mixture of ordered antiparallel hairpin and double-stranded helical structures in solutions of 0.1 or 0.5 M NaCl between 5 and 40 degrees C. The dimeric duplex is favored by conditions of high salt and low temperature. The monomer-dimer equilibrium constants vary from 5 x 10(6) to 5 x 10(3) and are unique for each DNA dodecamer. Analysis of the temperature dependence of the equilibrium constants shows that the double helix to hairpin conversion is driven by a positive entropy change and is associated with an endothermic enthalpy change. The mispair substitutions at the 5.8 positions and the IA(6.7) mispair have the greatest tendency toward hairpin formation and exhibit significantly larger entropy changes than the nonmispaired dGGTACGCGTACC parent sequence and the thermodynamically similar GA(6.7) DNA. The consequences of such hairpin-double helix equilibria must be considered in the interpretation of other kinds of experiments carried out on oligonucleotides at different concentrations.

Base Sequence↗
PubMed1991

Pre-existing fractures and bone mass predict vertebral fracture incidence in women.

OBJECTIVE: To determine the independent contributions of bone mass and existing fractures as predictors of the risk for new vertebral fractures. SUBJECTS: Postmenopausal Japanese-American women. MEASUREMENTS: Baseline measurements of the distal radius, the proximal radius, and the calcaneus were obtained in 1981 using single-photon absorptiometry. Measurements of the lumbar spine were obtained in 1984 using dual-photon absorptiometry. Prevalent vertebral fractures were identified using dimensions measured on lateral radiographs; vertebral height values more than 3 SD below vertebra-specific means were considered to indicate fracture. Statistical models were used to evaluate the utility of bone mass and existing (prevalent) fractures to predict the risk for new fractures during an average follow-up of 4.7 years. MAIN RESULTS: Differences of 2 SD in bone mass were associated with fourfold to sixfold increases in the risk for new vertebral fractures. A single fracture at the baseline examination increased the risk for new vertebral fractures fivefold. Presence of two or more fractures at baseline increased the risk 12-fold. A combination of low bone mass (below the 33d percentile) and the presence of two or more prevalent fractures increased the risk 75-fold, relative to women with the highest bone mass (above the 67th percentile) and no prevalent fractures. Stature, body mass index, arm span, and spinal conditions such as scoliosis, osteoarthritis, and sacroiliitis did not predict fracture incidence (P greater than 0.05). Weight was marginally predictive (P = 0.04) of fracture incidence but became nonpredictive after adjusting for bone mass (P greater than or equal to 0.05). CONCLUSIONS: Both bone mass and prevalent vertebral fractures are powerful predictors of the risk for new vertebral fractures. Combining information about bone mass and prevalent fracture appears to be better for predicting new fractures than either variable alone. Physicians can use these risk factors to identify patients at greatest risk for new fractures.

Absorptiometry, Photon↗
PubMed1991

A comparison of hip fracture incidence among native Japanese, Japanese Americans, and American Caucasians.

Hip fracture incidence rates among men and women of Japanese ancestry living on Oahu, Hawaii, from 1979 to 1981 were compared with rates for Japanese living on Okinawa, Japan, from 1984 to 1985. Both rates were further compared with those for various American Caucasian populations, including that of Rochester, Minnesota, from 1978 to 1982. Age-specific and cumulative (age 50-84 years) hip fracture rates among persons of Japanese ancestry were approximately half that of Caucasians for both sexes. Although diet and other cultural attributes of the Oahu group have become more westernized than the Okinawan population, there were no detectable differences in hip fracture rates between Oahu and Okinawa Japanese. Additional studies are needed to determine whether the observed differences in fracture rates between Caucasians and Japanese might be related to differences in bone mass, body size, frequency of falls, or other factors.

Adult↗
PubMed1991

Hairpin formation in the self-complementary dodecamer d-GGTACGCGTACC and derivatives containing GA and IA mispairs.

The dodecamer d-GGTACGCGTACC and four derivatives with GA and IA mispairs in the 6,7 and 5,8 positions have been examined in dilute solution and 0.01-0.1 M sodium chloride. Concentration dependence of Tm, gel electrophoresis, and equilibrium centrifugation indicate that these self-complementary oligomers can form hairpins under the present conditions. Thermal transitions measured in the ultraviolet primarily represent melting of hairpin to coil [cf. Scheffler et al. (1968, 1970)]. The Tm values show little or no depression for 6,7 substitution but rather large depression for 5,8 replacement. We interpret the results to indicate that the 6,7 sequences have two-base loops and five base pair stems and that the 5,8 sequences have four-base loops and four base pair stems. A concurrent theoretical modeling study [Raghunathan et al. (1991) Biochemistry (following paper in this issue)] provides support for this interpretation.

Base Composition↗
PubMed1991

Evaluation of adverse health outcomes associated with vertebral fractures.

Little is known about the frequency or degree to which vertebral fractures cause pain and physical disability. The purpose of this investigation was to examine the advantages of risk analysis over other statistical techniques (e.g., correlation analysis) for quantifying relationships between vertebral fractures and outcomes such as pain and disability. Subjects who volunteered to participate in studies of osteoporosis were asked about pain and disability. The number and degree of vertebral deformities were assessed from radiographs. Strong associations were observed between the most severe vertebral deformities and the risk of high pain or disability scores, while weaker associations were observed for moderate deformities. There did not appear to be any association between vertebral deformity and risk of moderate levels of pain or disability. Because of the potential for bias in cross-sectional studies such as this, the magnitude of these findings must be considered tentative. We conclude that risk analysis is an appropriate method for quantifying the relationship of vertebral fractures with pain and disability, but that prospective studies are now needed.

Back Pain↗
PubMed1991

Long-term precision of bone loss rate measurements among postmenopausal women.

Repeated measurements of bone mineral content can indicate the rate of bone loss among postmenopausal women. The clinical utility of such loss rate measurements will depend upon the long-term precision of the measurements. We have analyzed the precision of appendicular bone measurements among 495 Japanese-Americans followed for an average of 5.3 years and of both appendicular and axial measurements among 70 clinical trial participants followed for 2 years. Tables were derived from these analyses to quantitate the precision of individual loss rates under varying measurement conditions that might be encountered in clinical practice. The results demonstrate that only unusually rapid loss rates could be identified with confidence within short intervals, such as 1 year or 2. Extending the length of follow-up, however, appreciably improved the measured loss rate precision. In comparisons between bone sites, appendicular sites were determined to achieve a specified precision within the shortest intervals, followed by spine dual photon absorptiometry measurements. Spine quantitative computerized tomography measurements and measurements of hip sites required considerably longer follow-up intervals to achieve comparable precision.

Aged↗
PubMed1991

The clinical application of serial bone mass measurements.

Although the short-term precision of various bone mineral content (BMC) measurements is known, questions about the clinical use of serial BMC measurements remain: how frequently should BMC be measured? When is it appropriate to calculate bone loss rates? How are estimates of loss rate interpreted? This paper discusses both biological and technical sources of uncertainty, and the estimation of confidence limits for measured bone loss rates. For many, possibly most, patients, calculation of bone loss rate may not be necessary; however, repeated measures of BMC can still be useful for re-evaluating fracture risk. Indications for repeating BMC measurements may include low initial BMC (moderate to high fracture risk), anticipation of rapid bone loss (e.g., menopause, estrogen discontinuation), and verification of treatment efficacy.

Bone Density↗
PubMed1991

Age-related changes in bone mass among Japanese-American men.

Bone mass and appendicular bone loss rates were examined in a cohort of Japanese-American men. Across their age range (ages 61 to 82 years) bone mass steadily declined at the proximal and distal radius, and at the calcaneus. The cross-sectional reduction in bone mass was 3.5-6.3% per decade at the various sites. Longitudinal measurements of the same cohort indicated greater losses than suggested by the cross-sectional data, yet still less than 10% per decade. Linear trends of increasing loss rates with aging were significant at the calcaneus, and marginally significant at the radius sites. However, the oldest men in the cohort strongly influenced these trends. Men under age 75 had essentially constant annual rates of bone loss. The most elderly men had both the lowest bone mass and the greatest bone loss rates.

Age Factors↗
PubMed1991

Characteristics of respondents and nonrespondents in a prospective study of osteoporosis.

During 1981-1982, a cohort of elderly Japanese Americans living in Hawaii was recruited for an epidemiologic study of osteoporosis. The male subjects were simultaneously being examined for an epidemiologic study of heart disease. Baseline data collected from both the men and women at a previous heart disease examination were used to compare responders vs nonresponders. The target population for the osteoporosis study consisted of 1685 men and 1594 women. Of these, 1379 men (81.8%) and 1105 women (72.0%) participated in the initial osteoporosis examination. For each sex, nonrespondents were older and had higher systolic blood pressure levels than did the respondents. Male nonresponders had a higher stroke prevalence and more frequent recent use of vasodilator medicine. Female nonresponders had a less frequent history of having ever taken female hormones than did the responders. The responders and nonresponders were reasonably similar in other respects, as indicated by the comparison of more than 40 other variables. This suggests that nonresponse bias is probably not a major influence in exposure-disease associations in this osteoporosis cohort. We believe this is the first published report dealing with nonresponse characteristics in a cohort study of osteoporosis.

Aged↗
PubMed1991

Vertebral dimension differences between Caucasian populations, and between Caucasians and Japanese.

Various criteria have been proposed for using vertebral measurements to identify vertebral fractures. It is known that the normal distributions of vertebral heights and ratios vary with location within the spine. However, very little is known regarding the degree to which differences in these parameters may exist between populations. We report the vertebra-specific distributions of vertebral dimensions and ratios for Japanese-Americans, and compare these values to published data for Caucasians. The mean Japanese vertebral heights were 1 to 2 mm shorter than Caucasians, which may be due in part to the shorter stature of Japanese. However, differences in mean values were also observed between Caucasian populations. Furthermore, anterior/posterior vertebral height ratios differed between Caucasian studies, and between races. Additional studies are needed to determine to what degree these differences are due to technical and biological factors before criteria derived from one population can be used for identifying vertebral fractures in other populations of the same, or different, race.

Adult↗
PubMed1990

Ligand-induced biphasic protein denaturation.

The results of a thermodynamic calculation of the excess heat capacity that is based on experimental observations and that incorporates the effects of ligand binding on the two-state, thermal denaturation of a protein are presented. For a protein with a single-binding site on the native species and at subsaturating concentrations of ligand, bimodal or unimodal thermograms were computed merely by assuming a larger or smaller ligand association constant, respectively. The calculated thermograms for this simplified case show the salient features of those observed by differential scanning calorimetry for defatted human albumin monomer in the absence and presence of three ligands for which the protein has higher, intermediate, and lower affinity (Shrake, A., and Ross, P. D. (1988) J. Biol. Chem. 263, 15392-15399). The computation demonstrates that biphasic unfolding can result from a significant increase in the free energy of denaturation (and the transition temperature) during the course of unfolding due to a substantial increase in free ligand concentration caused by the release of bound ligand by denaturing protein. Such ligand-induced biphasic denaturation does not relate to macromolecular substructure but derives from a perturbation, during unfolding, of the ligand binding equilibrium, which is coupled to the equilibrium between the folded and unfolded protein species. Thus, this bimodality is not limited to thermally induced unfolding but is operative independent of the means used to effect denaturation and therefore must be considered when studying any macromolecular folding/unfolding reaction in the presence of ligand.

Kinetics↗