Residue lysine-34 in GroES modulates allosteric transitions in GroEL

The conserved residue Lys-34 in GroES was replaced by alanine and glutamic acid using site-directed mutagenesis. This residue is near the carboxy terminus of the mobile loop in GroES (residues 17-32) which becomes immobilized upon formation of the GroEL/GroES complex [Landry et al. (1993) Nature 364, 255-258]. Both charge neutralization (Lys-34-->Ala) and charge reversal (Lys-34-->Glu) at this position have little effect on the binding constant of GroES to GroEL, but they increase the enhancement by GroES of cooperativity in ATP hydrolysis by GroEL. This is reflected by a change in the Hill coefficient (at 10 mM K+) from 4.10 (+/- 0.22) in the presence of wild-type GroES to 5.17 (+/- 0.24) and 4.46 (+/- 0.14) in the presence of the GroES mutants Lys-34-->Ala and Lys-34-->Glu, respectively. The results are interpreted using the Monod-Wyman-Changeux (MWC) model for cooperativity [Monod et al. (1965) J. Mol. Biol. 12, 88-118]. They suggest that Lys-34 in GroES modulates the allosteric transition in GroEL by stabilizing a relaxed (R)-like state.     

DNA length and concentration dependencies of anisotropic phase transitions of DNA solutions

Critical concentrations for the isotropic to cholesteric phase transitions of double-stranded DNA fragments in simple buffered saline (0.1 M NaCl) solutions were determined as a function of DNA contour length ranging from approximately 50 nm to 2700 nm, by solid-state 31P NMR spectroscopy and polarized light microscopy. As expected for semirigid chains, the critical concentrations decrease sharply with increasing DNA length near the persistence length in the range from 50 to 110 nm, and approach a plateau when the contour length exceeds 190 nm. The biphasic region is substantially wider than observed for xanthan, another semirigid polyelectrolyte approximately twice as stiff as DNA, primarily because of low critical concentrations for first appearance of the anisotropic phase, C(i)*, in DNA samples > or =110 nm (320 base pairs) long. The limiting C(i)* for DNA > or =490 nm long is exceptionally low (only 13 mg/ml) and is substantially lower than the C(i)* of approximately 40 mg/ml reported for the stiffer xanthan polyelectrolyte. The much higher values of the critical concentrations, C(a)*, for the disappearance of the isotropic DNA phase (> or =67 mg/ml) are modestly higher than those observed for xanthan and are predicted reasonably well by a theory that has been applied to other semirigid polymers, if a DNA persistence length in the consensus range of 50-100 nm is assumed. By contrast, the broad biphasic region and low C(i)* values of DNA fragments > or =190 nm long could only be reconciled with theory by assuming persistence lengths of 200-400 nm. The latter discrepancies are presumed to reflect some combination of deficiencies in current theory as applied to chiral, strong polyelectrolytes such as DNA, and sequence-dependent variations in DNA properties such as flexibility, curvature, or interaction potential. The propensity of DNA to spontaneously self-order at low concentrations well in the physiological range may have biological significance.     

The question of neutrality in psychotherapy

The question of neutrality in psychotherapy is considered in the light of original historical psychoanalytic attitudes, and present usual practices. A disparity between what is reported and what is done is examined, and explanations for its existence are put forward.     

Ligand-induced conformational transitions in Escherichia coli phosphofructokinase 2: evidence for an allosteric site for MgATP2-

The binding of ligands to phosphofructokinase 2 (Pfk-2) from Escherichia coli induces changes in the fluorescence emission properties of its single tryptophan residue, Trp88, suggesting that upon binding the protein undergoes a conformational change. This fluorescence probe was used to determine the presence of an allosteric site for MgATP2- in the enzyme. Fructose 6-phosphate (fructose-6-P), the first substrate that binds to the enzyme with an ordered bi-bi mechanism, increases the fluorescence up to 30%. The saturation curve for this compound is hyperbolic with a Kd of 6 microM. The titration of Pfk-2 with MgATP2- causes a quenching of fluorescence of about 30% of its initial value, with a blue shift of 7 nm in the emission maximum. The response is cooperative with a Kd of 80 microM and a Hill coefficient of 2. The interaction of MgATP2- cannot take place at the active site in the absence of fructose-6-P, due to the ordered kinetic mechanism. Addition of compounds that bind to the catalytic site of Pfk-2, such as ATP4- or Mg-AMP-PNP, did not produce significant changes in the fluorescence spectrum of Trp88. However, in the absence of Mg2+, the addition of ATP4- to the enzyme-fructose-6-P complex shows a hyperbolic increase of fluorescence of 8%. Acrylamide steady-state quenching experiments for different enzyme-ligand complexes of Pfk-2, indicate that the tryptophan in the enzyme-MgATP2- complex is exposed to a much smaller extent to the solvent than in the free enzyme or in the enzyme-fructose-6-P complex. The effect of the binding of fructose-6-P or MgATP2- on the polarization fluorescence of the emission of Trp88 in Pfk-2 indicates changes in the local mobility of the Trp88 in both enzyme complexes. The average lifetime for Trp88 in Pfk-2 was found to be unusually large, approximately 7.7 ns, and did not vary significantly with the ligation state of the enzyme, which demonstrates that the quenching or enhancement of fluorescence induced by the ligands is mainly due to the complex formation with Pfk-2. These results demonstrate the presence of an allosteric site for MgATP2- in Pfk-2 from E. coli, responsible for the inhibition of the enzyme activity by this ligand.     

Two superhelix density-dependent DNA transitions detected by changes in DNA adsorption/desorption behavior

The adsorption behavior of covalently closed circular plasmid DNA at the mercury/water interface was studied by means of AC impedance measurements. The dependence of the differential capacitance (C) of the electrode double layer on the potential (E) was measured in the presence of adsorbed DNA. It was found that the C-E curves of supercoiled DNA at native and highly negative superhelix densities (sigma), relaxed covalently closed circular DNA, and nicked DNA differed from each other. A detailed study of topoisomer distributions ranging from -sigma of 0 to 0.11 revealed two supercoiling-dependent transitions, at about -sigma = 0.04 (transition TI) and 0.07 (transition TII). Transition TI was detected by measuring the height of the adsorption/desorption peak 1 (at about -1.2 V against the saturated calomel electrode) and the decrease of capacitance (DeltaC) at -0.35 V. This transition may be due to a sudden change in the ability of the DNA to respond to the alternating voltage, probably caused by changes in the DNA tertiary and/or secondary structure. Transition TII was detected by measuring peak 3* (at about -1.3 V), which was absent in topoisomers with -sigma less than 0.05. This transition is due to changes in the DNA adsorption/desorption behavior related to increased accessibility of bases at elevated negative superhelix density. Opening of the duplex at highly negative superhelix density was also detected by the single-strand selective probe of DNA structure, osmium tetroxide, 2, 2'-bipyridine. Our results suggest that electrochemical techniques provide sensitive experimental analysis of changes in DNA structure.     

Finite-size effects on long-range correlations: implications for analyzing DNA sequences

We analyze the fluctuations in the correlation exponents obtained for noncoding DNA sequences. We find prominent sample-to-sample variations as well as variations within a single sample in the scaling exponent. To determine if these fluctuations may result from finite system size, we generate correlated random sequences of comparable length and study the fluctuations in this control system. We find that the DNA exponent fluctuations are consistent with those obtained from the control sequences having long-range power-law correlations. Finally, we compare our exponents for the DNA sequences with the exponents obtained from power-spectrum analysis and correlation-function techniques, and demonstrate that the original "DNA-walk" method is intrinsically more accurate due to reduced noise.     

The God question in psychoanalysis.

This essay addresses certain complexities of dealing with the God-concept in psychoanalytic terms. Preanalytic philosophical and theological parallels to understanding the existence and nature of God find their echoes in psychoanalytic formulations of the God-concept. Centered on the idea of the God-representation, questions arise concerning the function of this representation as expressing the person's internal psychic reality as opposed to having some reference to a really existing divinity. Tensions in current analytic approaches to this problem are discussed, and suggestions are offered for advancing the potential dialogue in terms of the God-representation as a form of transitional conceptualization. Implications for the therapeutic handling of related issues are also suggested. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The epistemology question in psychology.

That psychologists are debating epistemological questions appears to be a relatively recent phenomenon only if one has no appreciation for the history of the discipline. Whether we evoke the writings of Wundt, Hull, Tolman, or more recent cognitive psychologists, it is impossible to escape the recurrence of certain fundamental questions on the nature of psychological knowledge and the justification of that knowledge. Even Skinner is a remarkably adept epistemologist despite his avowal that theories of learning are not necessary. But psychology has been enthralled for the better part of this century by a caricature of the very science it would become, and, while it has been consistently and severely criticized for this mistake, it has shown itself to be remarkably immune to such criticism. Part of the reluctance to change has stemmed from the lack of a serious indigenous alternative to the hackneyed version of logical empiricism and its operationist credo that have held sway over the discipline for the better part of its life. The papers included in the present issue of Canadian Psychology/Psychologie Canadienne are a small wedge into the broader conversation concerning what it means to have psychological knowledge. Each of these was part of a symposium presented at the Canadian Psychological Association Meetings in Montreal in 1988 by the Section on the History and Philosphy of Psychology. Each of the four papers argues (a) against the status quo of the shop-worn operationism and its attendant claims which pass for epistemology in psychology, and (b) for a renewed emphasis of either some one form of realism or constructionism. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The question cube: a model for developing question repertoire in training couple and family therapists

This paper presents a three-dimensional model for teaching questioning to those wishing to develop skills in couple and family therapy. The model breaks questions into their component parts of format (the style of the question: open, closed, forced choice, rating, or ranking); orientation (the person who is being inquired about: self or other), and subject (the content of the question: behavior, feelings, beliefs, meaning, or relationship). The model is presented in the context of our post-Milan version of couple and family therapy training. The model is useful in that it allows students gradually to increase their repertoire of questions in a way that offers step-wise learning and integrates with their existing skills.     

The question of lay analysis reconsidered.

Theodor Reik was central to the struggle for the legitimization of nonmedical or lay analysis both in Europe and the United States. This paper deals with the book Freud wrote in defense of Reik, The Question of Lay Analysis, and places it within its historical context. It then discusses the resistance to lay analysis by the New York Psychoanalytic Society and speculates on some of the reasons for this opposition. Finally, it discusses the state of nonmedical analysis today and a perspective for the future. (PsycINFO Database Record (c) 2010 APA, all rights reserved)