Allostery of 3-deoxy-D-arabino-heptulosonate 7-phosphate synthetase in Clostridium: another conserved generic characteristic

Enzymological studies were done to characterize the allosteric control of 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthetase in three species of Clostridium. Allosteric control was identified as feedback inhibition by phenylalanine and was qualitatively similar for the DAHP synthetases of C. butyricum, C. acetobutylicum, and C. tetanomorphum. Quantitative differences in the enzymology and kinetics of allosteric control distinguished C. tetanomorphum from C. butyricum and C. acetobutylicum. Crude extracts contained apparent proteolytic activity which could be fractionated from DAHP synthetase. The proteolytic activity was more labile than DAHP synthetase in extracts and was progressively inactivated by serial freeze-thaw treatments. Protease activity was at least partially inhibited by phenylmethylsulfonyl-fluoride. The method of comparative allostery of DAHP synthetase distinguishes the genera Bacillus and Clostridium, each having a strongly conserved pattern of regulation for DAHP synthetase. The data reinforce previous conclusions that allosteric control patterns governing the activity of DAHP synthetase are stable, reliable generic characteristics.     

Optimal design of feedback control by inhibition

Summary A recently developed, nonlinear method of systems analysis has been used to compare alternative patterns of control by feedback inhibition in otherwise equivalent unbranched biosynthetic pathways. The steady state performance of the simple case with end-product inhibition at the first physiologically important step is optimal with respect to the following criteria: the ability to (i) meet an increased demand for the end-product, (ii) meet this increased demand with limited accumulation of the intermediates, (iii) respond to an increased supply of the initial substrate, and (iv) limit the accumulation of the intermediates while responding to the increased availability of the initial substrate. The importance of these properties for the selection of feedback patterns in biosynthetic pathways is discussed.     

Regulation of the Biosynthesis of Amino Acids of the Aspartate Family in Coliform Bacteria and Pseudomonads

The control of aspartokinase and homoserine dehydrogenase activities was compared in aerobic and fermentative pseudomonads (genera Pseudomonas and Aeromonas), and in coliform bacteria representative of the principal genera of the Enterobacteriaceae. Isofunctional aspartokinases subject to independent end-product control occur in the Enterobacteriaceae and in Aeromonas. In Pseudomonas, there appears to be a single aspartokinase, subject to concerted feedback inhibition by lysine and threonine. Within this genus, the sensitivity of aspartokinase to the single allosteric inhibitors varies considerably: the aspartokinase of the acidovorans group is little affected by the single inhibitors, whereas that of the fluorescent group is severely inhibited by either amino acid at high concentration. In all bacteria examined, homoserine dehydrogenase activity is inhibited by threonine; inhibition is more severe in aerobic pseudomonads than in the other groups. In most of the bacteria examined, either nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate can serve as a cofactor for this enzyme, though the relative activity with the two pyridine nucleotides varies widely. Aerobic pseudomonads of the acidovorans group contain a homoserine dehydrogenase that is absolutely specific for NAD. The taxonomic implications of these findings are discussed.     

Effect of overall feedback inhibition in unbranched biosynthetic pathways

We have determined the effects of control by overall feedback inhibition on the systemic behavior of unbranched metabolic pathways with an arbitrary pattern of other feedback inhibitions by using a recently developed numerical generalization of Mathematically Controlled Comparisons, a method for comparing the function of alternative molecular designs. This method allows the rigorous determination of the changes in systemic properties that can be exclusively attributed to overall feedback inhibition. Analytical results show that the unbranched pathway can achieve the same steady-state flux, concentrations, and logarithmic gains with respect to changes in substrate, with or without overall feedback inhibition. The analytical approach also shows that control by overall feedback inhibition amplifies the regulation of flux by the demand for end product while attenuating the sensitivity of the concentrations to the same demand. This approach does not provide a clear answer regarding the effect of overall feedback inhibition on the robustness, stability, and transient time of the pathway. However, the generalized numerical method we have used does clarify the answers to these questions. On average, an unbranched pathway with control by overall feedback inhibition is less sensitive to perturbations in the values of the parameters that define the system. The difference in robustness can range from a few percent to fifty percent or more, depending on the length of the pathway and on the metabolite one considers. On average, overall feedback inhibition decreases the stability margins by a minimal amount (typically less than 5%). Finally, and again on average, stable systems with overall feedback inhibition respond faster to fluctuations in the metabolite concentrations. Taken together, these results show that control by overall feedback inhibition confers several functional advantages upon unbranched pathways. These advantages provide a rationale for the prevalence of this control mechanism in unbranched metabolic pathways in vivo.     

Intra- and intermolecular electron transfer processes in redox proteins

Initial velocity and product inhibition experiments were performed to characterize the kinetic mechanism of branched chain ketoacid dehydrogenase (the branched chain complex) activity. The results were directly compared to predicted patterns for a three-site ping-pong mechanism. Product inhibition experiments confirmed that NADH is competitive versus NAD+ and isovaleryl CoA is competitive versus CoA. Furthermore, both NADH and isovaleryl CoA were uncompetitive versus ketoisovaleric acid. These results are consistent with a ping-pong mechanism and are similar to pyruvate dehydrogenase and alpha-ketoglutarate dehydrogenase. However, inhibition patterns for isovaleryl CoA versus NAD+ and NADH versus CoA are not consistent with a ping-pong mechanism. These patterns may result from a steric interaction between the flavoprotein and transacetylase subunits of the complex. To determine the kinetic mechanism of the substrates and feedback inhibitors (NADH and isovaleryl CoA) of the branched chain complex, it was necessary to define the interaction of the inhibitors at nonsaturating fixed substrate (CoA and NAD+) concentrations. While the competitive inhibition patterns were maintained, slope replots for NADH versus NAD+ at nonsaturating CoA concentrations were parabolic. This unexpected finding resembles a linear mixed type of inhibition where the inhibition is a combination of pure competitive and noncompetitive inhibition.     

Biochemical diversity for biosynthesis of aromatic amino acids among the cyanobacteria.

We examined the enzymology and regulatory patterns of the aromatic amino acid pathway in 48 strains of cyanobacteria including representatives from each of the five major grouping. Extensive diversity was found in allosteric inhibition patterns of 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase, not only between the major groupings but also within several of the generic groupings. Unimetabolite inhibition by phenylalanine occurred in approximately half of the strains examined; in the other strains unimetabolite inhibition by tyrosine and cumulative, concerted, and additive patterns were found. The additive patterns suggest the presence of regulatory isozymes. Even though both arogenate and prephenate dehydrogenase activities were found in some strains, it seems clear that the arogenate pathway to tyrosine is a common trait that has been highly conserved among cyanobacteria. No arogenate dehydratase activities were found. In general, prephenate dehydratase activities were activated by tyrosine and inhibited by phenylalanine. Chorismate mutase, arogenate dehydrogenase, and shikimate dehydrogenase were nearly always unregulated. Most strains preferred NADP as the cofactor for the dehydrogenase activities. The diversity in the allosteric inhibition patterns for 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase, cofactor specificities, and the presence or absence of prephenate dehydrogenase activity allowed the separation of subgroupings within several of the form genera, namely, Synechococcus, Synechocystis, Anabaena, Nostoc, and Calothrix.     

A construct of interactive feedback control of the GH axis in the male

Growth hormone (GH) secretion is controlled by GH-releasing hormone (GHRH), the GH release-inhibiting hormone somatostatin (SRIF), and autofeedback connections. The ensemble network produces sexually dimorphic patterns of GH secretion. In an effort to formalize this system, we implemented a deterministically based autonomous feedback-driven construct of five principal dose-responsive regulatory interactions: GHRH drive of GH pituitary release, competitive inhibition of GH release by SRIF, GH autofeedback via SRIF with a time delay, delayed GH autonegative feedback on GHRH, and SRIF inhibition of GHRH secretion. This formulation engenders a malelike pattern of successive GH volleys due jointly to positive time-delayed feedback of GH on SRIF and negative feedback of SRIF on GH and GHRH. The multipeak volley is explicated as arising from a reciprocal interaction between GH and GHRH during periods of low SRIF secretion. The applicability of this formalism to neuroendocrine control is explored by initial parameter sensitivity analysis and is illustrated for selected feedback-dependent experimental paradigms. The present construct is not overparameterized and does not require an ad hoc pulse generator to achieve pulsatile GH output. Further evolution of interactive constructs could aid in exploring more complex feedback postulates that confer the vivid sexual dimorphism of female GH profiles.     

Quantitative control analysis of branched-chain 2-oxo acid dehydrogenase complex activity by feedback inhibition.

The potential for branched-chain 2-oxo acid dehydrogenase complex (BCOADC) activity to be controlled by feedback inhibition was investigated by calculating the Elasticity Coefficients for several feedback inhibitors. We suggest that feedback inhibition is a quantitatively important regulatory mechanism by which branched-chain 2-oxo acid dehydrogenase activity is regulated. The potential for control of enzyme activity is greater for NADH than for the acyl-CoA products, and suggests that factors that alter the redox potential may physiologically regulate BCOADC activity through a feedback inhibitory mechanism in vivo. Local pH may also be an important regulatory control factor.     

The effect of activation versus inhibition of feedback on perceived control of EEG activity

This study explores a model in which perceived control is affected by multiple sources of feedback at three different stages of the control sequence--person, response, and outcome. Behavior that enhances feedback is termed activation, while behavior that diminishes feedback is termed inhibition. The study tests the hypothesis that activation at any stage of the sequence leads to greater perceived control than inhibition. Eighty subjects increased or decreased their brain-wave activity (EEG) by making a tone go either on or off, using either an active or a passive strategy. Following 10 60-second trials, subjects rated their perception of control over their EEG activity. The hypothesis that making a tone go on (activation of the outcome) leads to a greater perception of control than making the tone go off (inhibition of the outcome) was confirmed only when subjects decreased their EEG activity. Perceived control was not significantly correlated with actual control, supporting the expectation that they are separately mediated. The results did not support the hypothesis that increasing EEG activity or using an activity strategy would lead to a greater perception of control than decreasing EEG or using a passive strategy. These findings are interpreted as evidence that attention to feedback may be necessary for the predicted bias in perceived control to occur, and that activation and inhibition should be operationalized as the absolute presence versus absence of feedback in testing the hypothesis for the first two stages of control.     

Feed-Forward versus Feedback Inhibition in a Basic Olfactory Circuit

Inhibitory interneurons play critical roles in shaping the firing patterns of principal neurons in many brain systems. Despite difference in the anatomy or functions of neuronal circuits containing inhibition, two basic motifs repeatedly emerge: feed-forward and feedback. In the locust, it was proposed that a subset of lateral horn interneurons (LHNs), provide feed-forward inhibition onto Kenyon cells (KCs) to maintain their sparse firing—a property critical for olfactory learning and memory. But recently it was established that a single inhibitory cell, the giant GABAergic neuron (GGN), is the main and perhaps sole source of inhibition in the mushroom body, and that inhibition from this cell is mediated by a feedback (FB) loop including KCs and the GGN. To clarify basic differences in the effects of feedback vs. feed-forward inhibition in circuit dynamics we here use a model of the locust olfactory system. We found both inhibitory motifs were able to maintain sparse KCs responses and provide optimal odor discrimination. However, we further found that only FB inhibition could create a phase response consistent with data recorded in vivo. These findings describe general rules for feed-forward versus feedback inhibition and suggest GGN is potentially capable of providing the primary source of inhibition to the KCs. A better understanding of how inhibitory motifs impact post-synaptic neuronal activity could be used to reveal unknown inhibitory structures within biological networks.     

Regular insulin secretory oscillations despite impaired ATP synthesis in Friedreich Ataxia patients.

Friedreich Ataxia is an inherited disorder caused by decreased expression of a mitochondrial protein called frataxin. Deficiency of this protein causes reduced biogenesis of iron-sulfur clusters, and subsequently impaired synthesis and replenishment of ATP IN VIVO. Basal secretion of insulin occurs in an oscillating manner presumably triggered by ATP-dependent feedback inhibition of glycolytic flux. Hence, individuals with reduced ATP synthesis rates should possibly exhibit impaired insulin secretory oscillations if these were solely dependent on ATP. In the present study Friedreich Ataxia patients with a presumptive impairment of ATP synthesis in pancreatic beta-cells were evaluated for regularity of basal secretory oscillations of insulin. Healthy siblings were employed as controls. In conflict with the initial hypothesis, no differences in regards to oscillation patterns were observed between patients and controls. Supported by EX VIVO evidence, these findings tentatively suggest that pulsatile insulin secretion might not be exclusively dependent on ATP feedback inhibition in humans.     

Role of feedback regulation of pantothenate kinase (CoaA) in control of coenzyme A levels in Escherichia coli

Pantothenate kinase (CoaA) is a key regulator of coenzyme A (CoA) biosynthesis in Escherichia coli, and its activity is controlled by feedback inhibition by CoA and its thioesters. The importance of feedback inhibition in the control of the intracellular CoA levels was tested by constructing three site-directed mutants of CoaA that were predicted to be feedback resistant based on the crystal structure of the CoaA-CoA binary complex. CoaA[R106A], CoaA[H177Q], and CoaA[F247V] were purified and shown to retain significant catalytic activity and be refractory to inhibition by CoA. CoaA[R106A] retained 50% of the catalytic activity of CoaA, whereas the CoaA[H177Q] and CoaA[F247V] mutants were less active. The importance of feedback control of CoaA to the intracellular CoA levels was assessed by expressing either CoaA or CoaA[R106A] in strain ANS3 [coaA15(Ts) panD2]. Cells expressing CoaA[R106A] had significantly higher levels of phosphorylated pantothenate-derived metabolites and CoA in vivo and excreted significantly more 4'-phosphopantetheine into the medium compared to cells expressing the wild-type protein. These data illustrate the key role of feedback regulation of pantothenate kinase in the control of intracellular CoA levels.     

The effect of activation versus inhibition of feedback on perceived control of EEG activity

This study explores a model in which perceived control is affected by multiple sources of feedback at three different stages of the control sequence — person, response, and outcome. Behavior that enhances feedback is termedactivation, while behavior that diminishes feedback is termedinhibition. The study tests the hypothesis that activation at any stage of the sequence leads to greater perceived control than inhibition. Eighty subjects increased or decreased their brain-wave activity (EEG) by making a tone go either on or off, using either an active or a passive strategy. Following 10 60-second trials, subjects rated their perception of control over their EEG activity. The hypothesis that making a tone go on (activation of the outcome) leads to a greater perception of control than making the tone go off (inhibition of the outcome) was confirmed only when subjects decreased their EEG activity. Perceived control was not significantly correlated with actual control, supporting the expectation that they are separately mediated. The results did not support the hypothesis that increasing EEG activity or using an activity strategy would lead to a greater perception of control than decreasing EEG or using a passive strategy. These findings are interpreted as evidence that attention to feedback may be necessary for the predicted bias in perceived control to occur, and that activation and inhibition should be operationalized as the absolute presence versus absence of feedback in testing the hypothesis for the first two stages of control.This article is based on a dissertation submitted to Yale University in partial fulfillment of the requirements for the doctoral degree.     

EMG feedback for the treatment of upper-extremity dysfunction: can it be effective?

This paper examines the application of EMG feedback for upper-extremity dysfunction secondary to neurologic injury. A rationale for the use of EMG feedback to enhance rotational components of upper-limb movement, train recruitment of the prime movers, and promote inhibition of motor responses that interfere with efficient and effortless movement is presented. Specific strategies that can be used to reinforce functional movement patterns are elaborated. A case study illustrating the application of the feedback strategies is provided. Despite sensory, perceptual, and cognitive impairments, a 53-year-old left hemiplegic obtained significant clinical upper-limb functional gains when given EMG feedback in conjunction with occupational therapy.     

下毛目纤毛虫的系统修订Ⅰ.原下毛亚目,排毛亚目(纤毛动物门)

对目前已发表下毛目纤毛虫的200个属级阶元进行了重新修订.给出新修订的下毛目系统,并根据其纤毛图式和形态发生特点将其归为5亚目20科82属.原下毛亚目Protohypotrichina suborder nov.,翁柯虫科Onychodromusidae fam.nov.,腹柱虫科Gastrostylidae fam.nov.和拟角毛虫属Parakeronopsis gen.nov.为新设,给出了下毛目中各亚目和科的特征简述及亚目及科的检索表.各属的特征以特征简述、检索表以及属的模式图的形式介绍之.新立原下毛亚目Protohypotrichina suborder nov.,并对排毛亚目Stichotrichina中的科属进行了重新修订.原下毛亚目的特征为无明确缘棘毛分化;排毛亚目的特征为无明确额腹横棘毛分化,腹棘毛为长的纵列,数目多,形式多样.修订后的原下毛亚目含2科3属,排毛亚目含5科29属.各科的基本特征分别为:原毛虫科Phacodinidae无明确体棘毛和背触毛的分化;凯毛虫科Kiitrichidae体纤毛器背腹有明确分化,有原始的背触毛分化;旋毛虫科Spirofilidae腹棘毛旋列或斜向排列;卡尔虫科Kahiellidae左、右缘棘毛均2至多列;小双虫科Amphisiellidae具1纵向排列的腹棘毛列;角毛虫科Keronopsidae具2列纵向排列的腹棘毛,通常无额棘毛及横棘毛的分化;翁柯虫科Onychodromusidae腹棘毛通常多列,一般有额、腹、横棘毛的初步分化;给出了各科及属的检索表,并附各属纤毛图式,对部分属进行讨论.     

Optimal design of feedback control by inhibition

The local stability of unbranched biosynthetic pathways is examined by mathematical analysis and computer simulation using a novel nonlinear formalism that appears to accurately describe biochemical systems. Four factors affecting the stability are examined: strength of feedback inhibition, equalization of the values among the corresponding kinetic parameters for the reactions of the pathway, pathway length, and alternative patterns of feedback interactions. The strength of inhibition and the pattern of feedback interactions are important determinants of steady-state behavior. The simple pattern of end-product inhibition in unbranched pathways may have evolved because it optimizes the steady-state behavior and is temporally most responsive to change. Stability in these simple systems is achieved by shortening pathway length either physically or, in the case of necessarily long pathways, kinetically by a wide devergence in the values of the corresponding kinetic parameters for the reactions of the pathway. These conclusions are discussed in the light of available experimental evidence.     

Biogeographical Interpretation of Elevational Patterns of Genus Diversity of Seed Plants in Nepal

This study tests if the biogeographical affinities of genera are relevant for explaining elevational plant diversity patterns in Nepal. We used simultaneous autoregressive (SAR) models to investigate the explanatory power of several predictors in explaining the diversity-elevation relationships shown in genera with different biogeographical affinities. Delta akaike information criterion (ΔAIC) was used for multi-model inferences and selections. Our results showed that both the total and tropical genus diversity peaked below the mid-point of the elevational gradient, whereas that of temperate genera had a nearly symmetrical, unimodal relationship with elevation. The proportion of temperate genera increased markedly with elevation, while that of tropical genera declined. Compared to tropical genera, temperate genera had wider elevational ranges and were observed at higher elevations. Water-related variables, rather than mid-domain effects (MDE), were the most significant predictors of elevational patterns of tropical genus diversity. The temperate genus diversity was influenced by energy availability, but only in quadratic terms of the models. Though climatic factors and mid-domain effects jointly explained most of the variation in the diversity of temperate genera with elevation, the former played stronger roles. Total genus diversity was most strongly influenced by climate and the floristic overlap of tropical and temperate floras, while the influences of mid-domain effects were relatively weak. The influences of water-related and energy-related variables may vary with biogeographical affinities. The elevational patterns may be most closely related to climatic factors, while MDE may somewhat modify the patterns. Caution is needed when investigating the causal factors underlying diversity patterns for large taxonomic groups composed of taxa of different biogeographical affinities. Right-skewed diversity-elevation patterns may be produced by the differential response of taxa with varying biogeographical affinities to climatic factors and MDE.     

Cephalic vasomotor feedback in the modification of migraine headache

The effect of cephalic vasomotor response (CVMR) and frontalis electromyographic (EMG) feedback on control of temporal arterial vasoconstriction and frontalis muscle activity in migraine and muscle contraction headache patients was investigated. A single subject multiple baseline design (across subjects and responses) was introduced to evaluate (1) patterning in the two physiological systems and (2) the effects of CVMR and EMG feedback on headache activity. The data indicated that (a) all four patients demonstrated an ability to control CVMR activity during CVMR feedback and EMG during EMG feedback, (b) idiosyncratic patterns of physiological activity emerge during feedback training, and (c) learned control of the pain mechanism for muscle contraction and migraine headaches was related to reduced frequency and duration of these headaches.Portions of this paper were presented at the Ninth Annual Convention of the Association for Advancement of Behavior Therapy, San Francisco, 1975.     

Regulation of the aromatic pathway in the cyanobacterium Synechococcus sp. strain Pcc6301 (Anacystis nidulans).

A pattern of allosteric control for aromatic biosynthesis in cyanobacteria relies upon early-pathway regulation as the major control point for the entire branched pathway. In Synechococcus sp. strain PCC6301 (Anacystis nidulans), two enzymes which form precursors for L-phenylalanine biosynthesis are subject to control by feedback inhibition. 3-Deoxy-D-arabino-heptulosonate 7-phosphate synthase (first pathway enzyme) is feedback inhibited by L-tyrosine, whereas prephenate dehydratase (enzyme step 9) is feedback inhibited by L-phenylalanine and allosterically activated by L-tyrosine. Mutants lacking feedback inhibition of prephenate dehydratase excreted relatively modest quantities of L-phenylalanine. In contrast, mutants deregulated in allosteric control of 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase excreted large quantities of L-phenylalanine (in addition to even greater quantities of L-tyrosine). Clearly, in the latter mutants, the elevated levels of prephenate must overwhelm the inhibition of prephenate dehydratase by L-phenylalanine, an effect assisted by increased intracellular L-tyrosine, an allosteric activator. The results show that early-pathway flow regulated in vivo by 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase is the dominating influence upon metabolite flow-through to L-phenylalanine. L-Tyrosine biosynthesis exemplifies such early-pathway control even more simply, since 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase is the sole regulatory enzyme subject to end-product control by L-tyrosine.