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W J Ray

Publications and source records attributed to W J Ray.

At least 37 records · Page 2Linked to original sources
PubMed1989

Comparison of rate constants for (PO3-) transfer by the Mg(II), Cd(II), and Li(I) forms of phosphoglucomutase.

Net rate constants that define the steady-state rate through a sequence of steps and the corresponding effective energy barriers for two (PO3-)-transfer steps in the phosphoglucomutase reaction were compared as a function of metal ion, M, where M = Mg2+ and Cd2+. These steps involve the reaction of either the 1-phosphate or the 6-phosphate of glucose 1,6-bisphosphate (Glc-P2) bound to the dephosphoenzyme (ED) to produce the phosphoenzyme (EP) and the free monophosphates, glucose 1-phosphate (Glc-1-P) or glucose 6-phosphate (Glc-6-P): EP.M + Glc-1-P----ED.M.Glc-P2----EP.M.Glc-6-P6. Before this comparison was made, net rate constants for the Cd2+ enzyme, obtained at high enzyme concentration via 31P NMR saturation-transfer studies [Post, C. B., Ray, W. J., Jr., & Gorenstein, D. G. (1989) Biochemistry (preceding paper in this issue)], were appropriately scaled by using the observed constants to calculate both the expected isotope-transfer rate at equilibrium and the steady-state rate under initial velocity conditions and comparing the calculated values with those measured in dilute solution. For the Mg2+ enzyme, narrow limits on possible values of the corresponding net rate constants were imposed on the basis of initial velocity rate constants for the forward and reverse directions plus values for the equilibrium distribution of central complexes, since direct measurement is not feasible. The effective energy barriers for both the Mg2+ and Cd2+ enzymes, calculated from the respective net rate constants, together with previously values for the equilibrium distribution of complexes in both enzymic systems [Ray, W. J., Jr., & Long, J. W. (1976) Biochemistry 15, 4018-4025], show that the 100-fold decrease in the kappa cat for the Cd2+ relative to the Mg2+ enzyme is caused by two factors: the increased stability of the intermediate bisphosphate complex and the decreased ability to cope with the phosphate ester involving the 1-hydroxyl group of the glucose ring. In fact, it is unlikely that the efficiency of (PO3-) transfer to the 6-hydroxyl group of bound Glc-1-P (thermodynamically favorable direction) is reduced by more than an order of magnitude in the Cd2+ enzyme. By contrast, the efficiency of the Li+ enzyme in the same (PO3-)-transfer step is less than 4 x 10(-8) that of the Mg2+ enzyme.(ABSTRACT TRUNCATED AT 400 WORDS)

Binding Sites↗
PubMed1987

The concentrations of free Mg2+ and free Zn2+ in equine blood plasma.

The enzyme phosphoglucomutase can be used as a metal ion indicator to measure the concentrations of free Mg2+ and free Zn2+ in physiological fluids. In horse plasma, the concentration of free Mg2+ is close to 0.5 mM, whereas that of free Zn2+ is about 2 X 10(-10) M, although numerous physiological roles for Zn2+ have been postulated that would require free Zn2+ concentration orders of magnitude higher than this. A titration of plasma with Zn2+ shows that the fractional increase in free Zn2+ is essentially the same as the fractional increase in total exchangeable Zn2+, and the results are consistent with a model in which essentially all of the Zn2+ in plasma is bound to albumin. Regardless of the model, the buffering capacity of plasma for free Zn2+ is intrinsically low; however, its capacity relative to the total (exchangeable) Zn2+ present is maximal. The implications of this type of buffering for homeostasis of plasma Zn2+ are considered. Treatment of plasma with a strong reducing agent such as dithiothreitol (0.1 mM) substantially increases the apparent binding of Zn2+ and thus reduces the free Zn2+ concentration. However, the concentration of free Zn2+ appears to be insensitive to decreases in the physiological concentrations of reduced glutathione and cysteine. The concentrations of free Zn2+ and free Mg2+ in plasma are similar to those that have been reported for muscle tissue (rabbit). Their ratio is about 4 X 10(-7). The physiological implications of these concentrations are considered. In some cases, if the Zn2+ and Mg2+ complexes of an uncharacterized vertebrate protein exhibit significantly different properties, their relative importance under physiological conditions can be approximated by evaluating those of the mixed complexes present in a solution that contains the physiological concentration of free Mg2+, plus Zn2+ buffered with histidine, at the appropriate pH and ionic strength. Other metal ion/chelon systems that come close to reproducing the concentrations of free Mg2+ and free Zn2+ in horse plasma also are considered.

Animals↗
PubMed1987

Remote nitrogen-15 isotope effects on addition of cyanide to NAD.

The reversible reaction NAD + CN(-)----NAD-CN was examined for remote secondary 15N isotope effects caused by isotopic substitution at the ring nitrogen of the nicotinamide group. These were compared with analogous effects for dehydrogenase-catalyzed reactions, since both cyanide and the hydride ion add at the N-4 position of the nicotinamide ring. The 15N effects on the rate constants for the forward and reverse processes were examined directly by conducting both the normal and isotopic reactions simultaneously under carefully controlled conditions in the sample and reference cells of a dual-beam spectrophotometer. In both cases, the 15N kinetic isotope effect differed from 1.00 by considerably less than 0.01. The 15N equilibrium isotope effect, 15K, was obtained as the ratio of equilibrium constants measured separately with natural-abundance and labeled NAD by using a concentration jump procedure [1.004 +/- 0.002 (cyanide addition)]. A similar value for 15K of 1.010 +/- 0.008 was obtained in an analogous manner for the reaction catalyzed by lactate dehydrogenase: NAD + lactate----pyruvate + NADH + H+. The latter value is significantly smaller than a previously reported value obtained from kinetic studies [1.044 +/- 0.012; Cook, P. F., Oppenheimer, N. J., & Cleland, W. W. (1981) Biochemistry 20, 1817]. The present value also is smaller than might be expected for a change in bond order from 4 to 3 [Cleland, W. W. (1980) Methods Enzymol. 64, 104-125] on the basis of the canonical resonance structures for NAD and NADH.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

The effect of polyethylene glycol on the growth and dissolution rates of a crystalline protein at high salt concentration. Phosphoglucomutase.

Although low concentrations of polyethylene glycol (1-5%, w/v) are essential for growing crystals of phosphoglucomutase from ammonium sulfate solutions (at close to 50% of saturation), the observed rate constant for short-term crystal growth on a defined, microcrystalline surface is essentially independent of polyethylene glycol concentration under these conditions. But this cosolute produces a substantial increase in the observed rate constant for the dissolution process and thus a corresponding increase in the solubility of the crystalline phase. These observations can be rationalized in terms of a decrease in the thermodynamic activity of the soluble form of phosphoglucomutase at high salt due to favorable interactions with polyethylene glycol (PEG) at the protein surface, coupled with a difference in accessibility of protein surfaces in the crystalline and solution states. Surfaces with a differential exposure in these two phases likely include both groups that interact favorably with polyethylene glycol relative to water (nonpolar groups) as well as those that interact unfavorably (ionic groups), but favorable PEG-protein interactions produced on dissolution must outweigh unfavorable ones. A PEG-induced increase in protein solubility at high salt concentration is likely to be general; PEG also may affect the growth of other protein crystals at high salt concentrations as it affects phosphoglucomutase.

Animals↗
PubMed1986

The structure of rabbit muscle phosphoglucomutase at intermediate resolution.

The three-dimensional structure of rabbit phosphoglucomutase has been determined to 2.7 A resolution by a combination of isomorphous and molecular replacement techniques. Heavy atom positions were found by using vector search and difference Fourier methods. The two molecules in the asymmetric unit form a dimer with its 2-fold axis perpendicular to and intersecting with a crystallographic 4(1) axis. Thus, the dimers are arranged so that they form fibers that are coincident with the 4(1) axes. A polypeptide model, corresponding with the known residue sequence, has been fitted to the electron density map to produce a structure that consists of four domains. All four have an alpha/beta structure; the first three have a somewhat similar topology that is based on a mixed parallel/antiparallel beta sheet, whereas the fourth is based on an antiparallel sheet. The active site lies between the four domains, with the phosphoserine residue in the first domain and some of the probable substrate-binding residues in the fourth and final domain. The carboxyl edges of all four sheets are directed towards the active site region, which lies in a deep crevice.

Animals↗
PubMed1986

The catalytic activity of muscle phosphoglucomutase in the crystalline phase.

A suspension of microcrystals of phosphoglucomutase in 60% ammonium sulfate exhibits a maximal catalytic activity in substrate-velocity studies that is about 0.2 of that obtained with the soluble enzyme under the same conditions. The apparent Michaelis constants for the reaction in the crystal phase are altered to an even smaller extent, relative to that in solution, although the parameters for the monophosphate and bisphosphate are increased more than 3 and more than 5 orders of magnitude, respectively, by the sulfate present. The compatibility of larger crystals with a reaction that constitutes part of the catalytic process also is demonstrated.

Animals↗
PubMed1986

Sex role and spatial ability: an EEG study.

The purpose of this study was to examine the relationship between sex role and spatial ability using both performance and electrocortical (EEG) measures. Two hundred and fifty four subjects were given the Differential Aptitudes Space Relations Test and a short form of the Personal Attributes Questionnaire (PAQ) which gives an indication of sex role. From this initial group, 40 males and females were chosen based on sex role scores. These subjects were asked to solve additional visuo-spatial problems. Bilateral EEG measures were taken from the frontal and parietal areas. The results indicate that frontal EEG measures are related to the sex role orientation of the subjects whereas parietal EEG measures are associated with performance on the visuo-spatial task for males but not females. These results suggest that frontal and parietal EEG measures reflect different processes. Counter to the traditional hypothesis, performance on the visuo-spatial task was negatively related to masculine sex-role orientation suggesting that factors which influence spatial processing go beyond biological sex.

Brain↗
PubMed1986

Microanatomy of the anterior cerebral artery.

The microanatomic features of the anterior cerebral artery were studied in 30 unfixed human brains which were injected with tinted polyester resin via cannulation of the internal carotid arteries under microscopic dissection. The outer diameter, length, and number of perforating branches were measured for each of the following vessels: anterior cerebral artery (proximal A1 segment, distal A2 segment), anterior communicating artery, and recurrent artery of Heubner. The perforating branches of the proximal segment of the anterior cerebral artery penetrated the brain at the anterior perforated substance, lateral chiasm, and optic tracts. The perforating branches of the anterior communicating artery penetrated the brain at the lamina terminalis, anterior perforated substance, and medial chiasm. The first 5 mm of the distal anterior cerebral artery (A2) had perforating branches penetrating the brain at the gyrus rectus and olfactory sulcus. The recurrent artery of Heubner originated from the A2 segment of the anterior cerebral artery in 57% of the cases, from the anterior cerebral artery-anterior communicating artery junction in 35%, and from the A1 segment in 8%. The depth of the interhemispheric fissure at the genu was 36.0 +/- 0.5 mm and at the midbody of the corpus callosum, 35.0 +/- 0.5 mm. Extension of the dissection to approach the anterior communicating artery from the genu of the corpus callosum using the anterior interhemispheric route was an additional 31.7 +/- 0.7 mm. The callosal arterial supply from the anterior cerebral artery showed short callosal branches in all brain specimens and long callosal vessels in 10% of the specimens.

Cerebral Arteries↗
PubMed1985

Active-site serine phosphate and histidine residues of phosphoglucomutase: pH titration studies monitored by 1H and 31P NMR spectroscopy.

1H and 31P NMR pH titrations were conducted to monitor changes in the environment and protonation state of the histidine residues and phosphoserine group of rabbit muscle phosphoglucomutase on binding of metal ions at the activating site and of substrate (glucose phosphate) at the catalytic site. Imidazole C epsilon-H signals from 8 of the 10 histidines present in the free enzyme were observed in 1H NMR spectra obtained by a spin-echo pulse sequence at 470 MHz; their pH (uncorrected pH meter reading of a 2H2O solution measured with a glass electrode standardized with H2O buffer) titration properties (in 99% 2H2O) were determined. Three of these histidine residues, which have pKa values ranging from 6.5 to 7.9, exhibited an atypical pH-dependent perturbation of their chemical shifts with a pHmid of 5.8 and a Hill coefficient of about 2. Since none of the observed histidines has a pKa near 5.8, it appears that these three histidines interact with a cluster consisting of two or more groups which become protonated cooperatively at this pH. Binding of Cd2+ at the activating site of the enzyme abolishes the pH-dependent transition of these histidines; hence, the putative anion cluster may constitute the metal ion binding site, or part of it. Two separate 31P NMR peaks from phosphoserine-116 of the phosphoenzyme were observed between pH 6 and 9. Apparently, the metal-free enzyme exists as a pH-dependent mixture of conformers that provide two different environments, I and II, for the enzymic phosphate group; the transition of the phosphate group between these two environments is slow on the NMR time scale.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

EEG alpha activity reflects attentional demands, and beta activity reflects emotional and cognitive processes.

Two experiments were designed to examine the effects of attentional demands on the electroencephalogram during cognitive and emotional tasks. We found an interaction of task with hemisphere as well as more overall parietal alpha for tasks not requiring attention to the environment, such as mental arithmetic, than for those requiring such attention. Differential hemispheric activation for beta was found most strongly in the temporal areas for emotionally positive or negative tasks and in the parietal areas for cognitive tasks.

Alpha Rhythm↗
PubMed1985

Multinuclear magnetic resonance studies of metal ion binding sites of phosphoglucomutase.

Metal binding at the activating site of rabbit muscle phosphoglucomutase has been studied by 31P, 7Li, and 113Cd NMR spectroscopy. A 7Li NMR signal of the binary Li+ complex of the phosphoenzyme was not observed probably because of rapid transverse relaxation of the bound ion due to chemical exchange with free Li+. The phosphoenzyme-Li+-glucose 6-phosphate ternary complex is more stable, kinetically, and yields a well-resolved peak from bound Li+ at -0.24 ppm from LiCl with a line width of 5 Hz and a T1 relaxation time of 0.51 +/- 0.07 s at 78 MHz. When glucose 1-phosphate was bound, instead, the chemical shift of bound 7Li+ was -0.13 ppm; and in the Li+ complex of the dephosphoenzyme and glucose bisphosphate a partially broadened 7Li+ peak appeared at -0.08 ppm. Thus, the bound metal ion has a somewhat different environment in each of these three ternary complexes. The 113Cd NMR signal of the binary Cd2+ complex of the phosphoenzyme appears at 22 ppm relative to Cd(ClO4)2 with a line width of 20 Hz at 44.4 MHz. Binding of substrate and formation of the Cd2+ complex of the dephosphoenzyme and glucose bisphosphate broaden the 113Cd NMR signal to 70 Hz and shift it to 75 ppm. The 53 ppm downfield shift upon the addition of substrate along with 1H NMR data suggests that one oxygen ligand to Cd2+ in the binary complex is replaced by a nitrogen ligand at some intermediate point in the enzymic reaction.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

A simple procedure for removing contaminating aldehydes and peroxides from aqueous solutions of polyethylene glycols and of nonionic detergents that are based on the polyoxyethylene linkage.

Commercial samples of polyethylene glycol as well as detergents incorporating the polyoxyethylene linkage may contain substantial concentrations of peroxides and aldehydes, as well as aldehyde precursors. Facile procedures for essentially eliminating either the first or all three of these contaminants from concentrated aqueous stock solutions are described. Under some conditions, the purified solutions can be stored at room temperature under aerobic conditions for time periods of weeks without significant decomposition. Possible antioxidants that might be used under other conditions to stabilize such solutions during long-term aerobic studies are considered.

Aldehydes↗
PubMed1985

EEG correlates of emotional tasks related to attentional demands.

This research brings together two separate areas: that of EEG processes associated with positive and negatively valenced emotional material; and that of traditional psychophysiological research related to the "intake" and "rejection" of environmental stimuli. Forty males on each of two days were presented with tasks reflecting both attentional demands and affectual processing. Heart rate and bilateral EEG measures from frontal, parietal and temporal sites were recorded. Using a FFT (fast Fourier transform) electrocortical activity in the 2-7 Hz, 8-15 Hz, and 16-24 Hz was determined and analyzed. The results suggest emotional valence (i.e. positive and negative) and attentional demands (i.e. intake vs rejection) are differentially represented in terms of EEG functioning. An interaction of attentional demand with hemisphere was found for EEG alpha activity in the temporal and parietal areas. For emotional valence there was a significant main effect for EEG beta activity in both the temporal and parietal areas. Differential hemispheric activity was found using a factor analytic technique (PARAFAC) with positively valenced tasks being associated with right temporal beta. Heart rate changes for the attentional dimension were consistent with previous research.

Adolescent↗
PubMed1985

EEG activity during cognitive processing: influence of attentional factors.

The research draws upon two separate areas of inquiry: that of lateralized electrocorical processing of cognitive material; and that of psychophysiological information processing studies related to foci of attentional demands. Eighteen subjects on each of three separate days were presented with tasks considered to be 'right hemispheric' or 'left hemispheric'. These tasks were paired in a 2 X 2 design with an attentional factor requiring attention to the environment or to internal processing. All subjects received all types of tasks. Bilateral EEG measures were taken from the frontal and parietal areas referenced to linked ears. Major frequency bands were determined by a Fast Fourier Transform (FFT) and appropriate ANOVAs performed. The results suggest that task factors (left vs right hemisphere tasks) and attentional demands (internal vs external) are differentially represented in terms of EEG functioning. In general the higher beta frequencies were more sensitive to the hemispheric tasks demands and the middle frequencies (alpha and low beta) more sensitive to the attentional demands especially in the parietal areas.

Alpha Rhythm↗
PubMed1984

Acceleration of the NAD cyanide adduct reaction by lactate dehydrogenase: the equilibrium binding effect as a measure of the activation of bound NAD.

The binary complex of NAD and lactate dehydrogenase reacts reversibly with cyanide to produce a complex (E X NAD-CN) whose noncovalent interactions are similar to those in the E X NADH complex (where E is one-fourth of the tetrameric dehydrogenase). The reaction apparently is a simple bimolecular nucleophilic addition at the 4 position of the bound nicotinamide ring; viz., cyanide does not bind to the enzyme prior to reaction. The value of the dissociation constant for E X NAD-CN is about 1 X 10(-6) M and is independent of pH over the range of 6-8. The equilibrium constant for the reaction of cyanide with E X NAD is about 400-fold larger than that for the nonenzymic process after a statistical correction. This increment in Ke is accounted for by a 220-fold increase in the rate of the forward enzymic reaction (20 M-1 s-1) as compared with an approximately 2-fold decrease for the reverse process (9 X 10(-5) s-1). Thus, the increased value of the rate constant for bond formation in the enzymic reaction is attributed to an equilibrium binding effect that is translated almost entirely into a rate effect on that step (bond formation). Since the nonenzymic reaction is sensitive to solvent composition, this equilibrium binding effect likely is produced by environmental effects at the nicotinamide/dehydronicotinamide part of the coenzyme binding site on the enzyme.

Animals↗
PubMed1984

The lactate dehydrogenase catalyzed pyruvate adduct reaction: simultaneous general acid-base catalysis involving an enzyme and an external catalyst.

The pH dependence of the reaction catalyzed by lactate dehydrogenase, where pyruvate adds covalently to NAD to yield a NAD-Pyr adduct, together with published data on the pH dependence of parameters in the normal redox reaction suggests similar binding modes for enolpyruvate and lactate in their complexes with E X NAD (where E is one-fourth of the tetramer), for ketopyruvate in its complexes with the protonated species, E X H X NAD and E X H X NADH, and for the NAD--Pyr adduct and NADH plus pyruvate in their complexes with E X H. These similarities, together with previous data, suggest a reaction scheme for the formation of the enzyme-adduct complex that includes the relevant proton-transfer steps. Seven different amine chloride buffers were used in a study of the reverse adduct reaction, i.e., the decomposition of E X H X NAD--Pyr. These act with varying efficiencies as external general acid catalysts; the enzyme apparently acts as a (internal) general base. The involvement of the amine chloride buffers as external general catalysts is supported by the concentration dependence of the buffer effect, by a Brönsted plot, and by solvent deuterium isotope effects. The involvement of the enzyme as an internal general catalyst is inferred from the pH dependence of the reaction and the identities of the nearby groups in the E X H X NAD--Pyr complex (from crystallographic studies). The dependence of the adduct reaction on chloride concentration indicates the presence of dead-end inhibitor complexes of E X H X Cl and E X H X NAD X Cl. Chloride also accelerates the decomposition of the adduct in the complex E X H X NAD--Pyr by binding to this complex.

Animals↗