A key question in understanding microtubule dynamics is how GTP hydrolysis leads to catastrophe, the switch from slow growth to rapid shrinkage. We first provide a review of the experimental and modeling literature, and then present a new model of microtubule dynamics. We demonstrate that vectorial, random, and coupled hydrolysis mechanisms are not consistent with the dependence of catastrophe on tubulin concentration and show that, although single-protofilament models can explain many features of dynamics, they do not describe catastrophe (...) as a multistep process. Finally, we present a new combined (coupled plus random hydrolysis) multiple-protofilament model that is a simple, analytically solvable generalization of a single-protofilament model. This model accounts for the observed lifetimes of growing microtubules, the delay to catastrophe following dilution and describes catastrophe as a multistep process. (shrink)

The translocation of tRNAs through the ribosome proceeds through numerous small steps in which tRNAs gradually shift their positions on the small and large ribosomal subunits. The most urgent questions are: (i) whether these intermediates are important; (ii) how the ribosomal translocase, the GTPase elongation factor G (EF‐G), promotes directed movement; and (iii) how the energy of GTP hydrolysis is coupled to movement. In the light of recent advances in biophysical and structural studies, we argue that intermediate states of (...) translocation are snapshots of dynamic fluctuations that guide the movement. In contrast to current models of stepwise translocation, kinetic evidence shows that the tRNAs move synchronously on the two ribosomal subunits in a rapid reaction orchestrated by EF‐G and GTP hydrolysis. EF‐G combines the energy regimes of a GTPase and a motor protein and facilitates tRNA movement by a combination of directed Brownian ratchet and power stroke mechanisms. (shrink)

Recently, Gao et al. and Chappie et al. elucidated the crystal structures of the polytetrameric stalk domain of the dynamin‐like virus resistance protein, MxA, and of the G‐domain dimer of the large, membrane‐deforming GTPase, dynamin, respectively. Combined, they provide a hypothetical oligomeric structure for the complete dynamin protein. Here, it is discussed how the oligomers are expected to form and how they participate in dynamin mediated vesicle fission during the process of endocytosis. The proposed oligomeric structure is compared with the (...) novel mechanochemical model of dynamin function recently proposed by Bashkirov et al. and Pucadyil and Schmid. In conclusion, the new model of the dynamin oligomer has the potential to explain how short self‐limiting fissogenic dynamin assemblies are formed and how concerted GTP hydrolysis is achieved. The oligomerisation of two other dynamin superfamily proteins, the guanylate binding proteins (GBPs) and the immunity‐related GTPases (IRGs), is addressed briefly. (shrink)

Microtubules form a highly dynamic filament network in all eukaryotic cells. Individual microtubules grow by tubulin dimer subunit addition and frequently switch between phases of growth and shortening. These unique dynamics are powered by GTP hydrolysis and drive microtubule network remodeling, which is central to eukaryotic cell biology and morphogenesis. Yet, our knowledge of the molecular events at growing microtubule ends remains incomplete. Here, recent ultrastructural, biochemical and cell biological data are integrated to develop a realistic model of growing (...) microtubule ends comprised of structurally distinct but biochemically overlapping zones. Proteins that recognize microtubule lattice conformations associated with specific tubulin guanosine nucleotide states may independently control major structural transitions at growing microtubule ends. A model is proposed in which tubulin dimer addition and subsequent closure of the MT wall are optimized in cells to achieve rapid physiological microtubule growth. (shrink)

Dynamin is a GTPase that regulates late events in clathrin‐coated vesicle formation. Our current working model suggests that dynamin is targeted to coated pits in its unoccupied or GDP‐bound form, where it is initially distributed uniformly throughout the clathrin lattice. GTP/GDP exchange triggers its release from these sites and its assembly into short helices that encircle the necks of invaginated coated pits like a collar. GTP hydrolysis, which is required for vesicle detachment, presumably induces a concerted conformation change, tightening (...) the collar. Unlike most of its GTPase cousins that serve as molecular switches, dynamin has a low affinity for GTP, a very high intrinsic rate of GTP hydrolysis and functions as a homo‐oligomer. A concerted conformational change resulting from coordinated GTP hydrolysis by the dynamin oligomer might be sufficient to generate force. In this case, dynamin would be the first GTPase identified that acts as a structural protein with mechano‐chemical function. (shrink)

The eukaryotic cytoskeleton appears to have evolved from ancestral precursors related to prokaryotic FtsZ and MreB. FtsZ and MreB show 40–50% sequence identity across different bacterial and archaeal species. Here I suggest that this represents the limit of divergence that is consistent with maintaining their functions for cytokinesis and cell shape. Previous analyses have noted that tubulin and actin are highly conserved across eukaryotic species, but so divergent from their prokaryotic relatives as to be hardly recognizable from sequence comparisons. One (...) suggestion for this extreme divergence of tubulin and actin is that it occurred as they evolved very different functions from FtsZ and MreB. I will present new arguments favoring this suggestion, and speculate on pathways. Moreover, the extreme conservation of tubulin and actin across eukaryotic species is not due to an intrinsic lack of variability, but is attributed to their acquisition of elaborate mechanisms for assembly dynamics and their interactions with multiple motor and binding proteins. A new structure‐based sequence alignment identifies amino acids that are conserved from FtsZ to tubulins. The highly conserved amino acids are not those forming the subunit core or protofilament interface, but those involved in binding and hydrolysis of GTP. BioEssays 29:668–677, 2007. © 2007 Wiley Periodicals, Inc. (shrink)

Rho/Rac proteins constitute a subgroup of the Ras superfamily of GTP hydrolases. Although originally implicated in the control of cytoskeletal events, it is currently known that these GTPases coordinate diverse cellular functions, including cell polarity, vesicular trafficking, the cell cycle and transcriptomal dynamics. In this review, we will provide an overview on the recent advances in this field regarding the mechanism of regulation and signaling, and the roles in vivo of this important GTPase family. BioEssays 29:356–370, 2007. © 2007 Wiley (...) Periodicals, Inc. (shrink)

Almost 40 years since the discovery of microtubule dynamic instability, the molecular mechanisms underlying microtubule dynamics remain an area of intense research interest. The “standard model” of microtubule dynamics implicates a “cap” of GTP‐bound tubulin dimers at the growing microtubule end as the main determinant of microtubule stability. Loss of the GTP‐cap leads to microtubule “catastrophe,” a switch‐like transition from microtubule growth to shrinkage. However, recent studies, using biochemical in vitro reconstitution, cryo‐EM, and computational modeling approaches, challenge the simple GTP‐cap (...) model. Instead, a new perspective on the mechanisms of microtubule dynamics is emerging. In this view, highly dynamic transitions between different structural conformations of the growing microtubule end – which may or may not be directly linked to the nucleotide content at the microtubule end – ultimately drive microtubule catastrophe. (shrink)

This article examines the political controversy in the United States surrounding a new process for the disposition of human remains, alkaline hydrolysis. AH technologies use a heated solution of water and strong alkali to dissolve tissues, yielding an effluent that can be disposed through municipal sewer systems, and brittle bone matter that can be dried, crushed, and returned to the decedent’s family. Though AH is legal in eight US states, opposition to the technology remains strong. Opponents express concerns about (...) public health and safety and about the dignity of our mortal remains. Proponents focus on AH’s environmental benefits over cremation and earth burial, aligning the technology with the “green burial” movement. Drawing from historical sources, Science, Technology, and Society literature, interviews with funeral professionals, industry literature, and various media sources, this article examines four prominent conceptions of the dead human body as they are deployed by various funeral stakeholders seeking to exercise authority over the dead human body, to influence the trajectory of AH technology in the United States, and to chart a course for US deathcare culture in the twenty-first century. (shrink)

This article morally assesses alkaline hydrolysis as a means of final bodily disposition. Arguing from the Catholic social and theological principles of human dignity, the doctrine of bodily resurrection, subsidiarity, and the common good, the author shows that, while alkaline hydrolysis has some advantages over burial and cremation, Catholic conferences should be encouraged to resist its legalization, provided they focus renewed energy on teaching the faithful about the significance of Christ’s victory, by the Resurrection, over the corruption of (...) bodily death. (shrink)

The ring‐shaped ATPase machine, cohesin, regulates sister chromatid cohesion, transcription, and DNA repair by topologically entrapping DNA. Here, we propose a rigid scaffold model to explain how the cohesin regulators Pds5 and Wapl release cohesin from chromosomes. Recent studies have established the Smc3‐Scc1 interface as the DNA exit gate of cohesin, revealed a requirement for ATP hydrolysis in ring opening, suggested regulation of the cohesin ATPase activity by DNA and Smc3 acetylation, and provided insights into how Pds5 and Wapl (...) open this exit gate. We hypothesize that Pds5, Wapl, and SA1/2 form a rigid scaffold that docks on Scc1 and anchors the N‐terminal domain of Scc1 (Scc1N) to the Smc1 ATPase head. Relative movements between the Smc1‐3 ATPase heads driven by ATP and Wapl disrupt the Smc3‐Scc1 interface. Pds5 binds the dissociated Scc1N and prolongs this open state of cohesin, releasing DNA. We review the evidence supporting this model and suggest experiments that can further test its key principles. (shrink)

Resistance to inhibitors of cholinesterase 8A (Ric‐8A) is a prominent non‐receptor GEF and a chaperone of G protein α‐subunits (Gα). Recent studies shed light on the structure of Ric‐8A, providing insights into the mechanisms underlying its interaction with Gα. Ric‐8A is composed of a core armadillo‐like domain and a flexible C‐terminal tail. Interaction of a conserved concave surface of its core domain with the Gα C‐terminus appears to mediate formation of the initial Ric‐8A/GαGDP intermediate, followed by the formation of a (...) stable nucleotide‐free complex. The latter event involves a large‐scale dislocation of the Gα α5‐helix that produces an extensive primary interface and disrupts the nucleotide‐binding site of Gα. The distal portion of the C‐terminal tail of Ric‐8A forms a smaller secondary interface, which ostensibly binds the switch II region of Gα, facilitating binding of GTP. The two‐site Gα interface of Ric‐8A is distinct from that of GPCRs, and might have evolved to support the chaperone function of Ric‐8A. (shrink)

The photocycle of visual opsins is essential to maintain the light sensitivity of the retina. The early physical observations of the rhodopsin photocycle by Böll and Kühne in the 1870s inspired over a century's worth of investigations on rhodopsin biochemistry. A single photon isomerizes the Schiff‐base linked 11‐cis‐retinylidene chromophore of rhodopsin, converting it to the all‐trans agonist to elicit phototransduction through photoactivated rhodopsin (Rho*). Schiff base hydrolysis of the agonist is a key step in the photocycle, not only diminishing (...) ongoing phototransduction but also allowing for entry and binding of fresh 11‐cis chromophore to regenerate the rhodopsin pigment and maintain light sensitivity. Many challenges have been encountered in measuring the rate of this hydrolysis, but recent advancements have facilitated studies of the hydrolysis within the native membrane environment of rhodopsin. These techniques can now be applied to study hydrolysis of agonist in other opsin proteins that mediate phototransduction or chromophore turnover. In this review, we discuss the progress that has been made in characterizing the rhodopsin photocycle and the journey to characterize the hydrolysis of its all‐trans‐retinylidene agonist. (shrink)

Release of the ATP hydrolysis product ortophosphate (Pi) from the active site of myosin is central in chemo‐mechanical energy transduction and closely associated with the main force‐generating structural change, the power‐stroke. Despite intense investigations, the relative timing between Pi‐release and the power‐stroke remains poorly understood. This hampers in depth understanding of force production by myosin in health and disease and our understanding of myosin‐active drugs. Since the 1990s and up to today, models that incorporate the Pi‐release either distinctly before (...) or after the power‐stroke, in unbranched kinetic schemes, have dominated the literature. However, in recent years, alternative models have emerged to explain apparently contradictory findings. Here, we first compare and critically analyze three influential alternative models proposed previously. These are either characterized by a branched kinetic scheme or by partial uncoupling of Pi‐release and the power‐stroke. Finally, we suggest critical tests of the models aiming for a unified picture. (shrink)

Summary In this contribution, I discuss the relevance of epistemological models for psychotherapy. Despite its importance epistemology is seldom explicitly dealt with in the psychotherapeutic landscape. Based on the presentation of “Critical Realism (CR),” the epistemological position of Gestalt Theoretical Psychotherapy (GTP), I intend to show to which extent this explanatory model supports a differentiated understanding of problems between human beings, arising from the differences in experiencing “reality.” The presentation deals explicitly with some conclusions that can be drawn from the (...) CR model for practical psychotherapeutic work. In particular, the aspects of basic therapeutic attitude, therapeutic relationship, and praxeology are highlighted. (shrink)

This study examines the hydrolysis of lignocellulose extracted from poultry manure for the purpose of investigating low-cost feedstocks for ethanol production while providing an alternative solid waste management strategy for agricultural livestock manures. Poultry manure underwent various pretreatments to enhance subsequent enzymatic hydrolysis including untreated, alkaline pretreatment with 0.5N KOH, drying, and grinding. The KOH-treated, dried, and grinded poultry manure yielded the highest glucose conversions. When poultry manure without pretreatment was hydrolyzed at 40°C with an enzyme loading 400 (...) units/g feedstock, 7.1 ± 0.3% was converted to glucose in 24 hours. This increased to 27.6 ± 1.2% when a KOH pretreatment followed by drying and grinding was applied. Fourier transform infrared spectroscopy analyses were used to examine differences in the composition of the pretreated feedstock before and after enzymatic hydrolysis. These indicate that the cellulose content in pretreated poultry manure was removed during enzymatic hydrolysis. (shrink)

Summary With regard to the dynamics of human experience and behavior, Gestalt theoretical psychotherapy (GTP) relies mainly on Kurt Lewin’s dynamic field theory of personality. GTP is carried out by including a re-interpretation of Lewin’s theory in some aspects of psychotherapeutic practice in relation to critical realism. Human experience and behavior are understood to be functions of the person and the environment (including the other individuals therein) in a psychic field (life space), which encompasses both of these mutually dependent factors. (...) The anthropological model of this approach is, therefore, not mono-personal but, a priori, structural and relational in nature. It does not one-sidedly focus on the “inner components” of a person, but on the interrelation of the individual and a given environment, which affects experience and behavior. After a brief introduction of these basic concepts, this lecture will focus especially on Lewin’s concept of tension-systems, which may be considered as the Gestalt theoretical counterpart of Freud’s drive theory. Further, we define the basic assumptions which underlie GTP and explain how the person moves through her/his life experience in terms of Gestalt psychology. (shrink)

The study of the effects of interactive media has mainly focused on dysregulated behaviors, the conceptualization of which is supported by the paradigms of addiction. Research into Game Transfer Phenomena examines the interplay between video game features, events while playing, and the manipulation of hardware, which can lead to sensory-perceptual and cognitive intrusions and self-agency transient changes related to video games. GTP can influence the interpretation of stimuli and everyday interactions and, in contrast to gaming disorder, are relatively common and (...) not necessarily negative. However, some players have reported feeling distress due to their GTP. This study focuses on how dispositional and interactive media habit factors are related to GTP and two forms of problematic interactive media [problematic video game playing and problematic social media use ]. A sample of 343 university students who played video games completed an online survey. Not all who had experienced GTP were identified as exhibiting PVG or PSMU, but all of those in the PVG group had experienced GTP. Overall, the profiles of the groups, including GTP, PVG, and PSMU, were in accordance with previous findings. Those in the GTP and the PVG groups were characterized by being male, being highly engaged in the game, and showed preferences for game-related activities. However, while those in the GTP group were significantly more likely to be fantasy-prone, those with PVG were the ones who played most per day. Those in the PSMU group were characterized by being female and/or extroverted, frequently using social/sharing platforms, and seldom playing video games. A hierarchical binary logistic regression revealed that males were more likely to experience GTP. Increases in PVG, fantasy proneness, and neuroticism increased the odds of GTP. Future work can benefit from considering the role of GTP in gaming disorder, since intrusive thoughts, cognitive biases, and poor impulse control are pivotal in the initiation and maintenance of dysfunctional playing behaviors. (shrink)

Intermediary metabolism molecules are orchestrated into logical pathways stemming from history (L-amino acids, D-sugars) and dynamic constraints (hydrolysis of pyrophosphate or amide groups is the driving force of anabolism). Beside essential metabolites, numerous variants derive from programmed or accidental changes. Broken down, variants enter standard pathways, producing further variants. Macromolecule modification alters enzyme reactions specificity. Metabolism conform thermodynamic laws, precluding strict accuracy. Hence, for each regular pathway, a wealth of variants inputs and produces metabolites that are similar to but (...) not the exact replicas of core metabolites. As corollary, a shadow, paralogous metabolism, is associated to standard metabolism. We focus on a logic of paralogous metabolism based on diversion of the core metabolic mimics into pathways where they are modified to minimize their input in the core pathways where they create havoc. We propose that a significant proportion of paralogues of well-characterized enzymes have evolved as the natural way to cope with paralogous metabolites. A second type of denouement uses a process where protecting/deprotecting unwanted metabolites - conceptually similar to the procedure used in the laboratory of an organic chemist - is used to enter a completely new catabolic pathway. (shrink)

Ran is one of the most abundant and best conserved of the small GTP binding and hydrolyzing proteins of eukaryotes. It is located predominantly in cell nuclei. Ran is a member of the Ras family of GTPases, which includes the Ras and Ras‐like proteins that regulate cell growth and division, the Rho and Rac proteins that regulate cytoskeletal organization and the Rab proteins that regulate vesicular sorting. Ran differs most obviously from other members of the Ras family in both its (...) nuclear localization, and its lack of sites required for post‐translational lipid modification. Ran is, however, similar to other Ras family members in requiring a specific guanine nucleotide exchange factor (GEF) and a specific GTPase activating protein (GAP) as stimulators of overall GTPase activity. In this review, the multiple cellular functions of Ran are evaluated with respect to its known biochemistry and molecular interactions. (shrink)

Protein α-helices provide an ordered biological environment that is conducive to soliton-assisted energy transport. The nonlinear interaction between amide I excitons and phonon deformations induced in the hydrogen-bonded lattice of peptide groups leads to self-trapping of the amide I energy, thereby creating a localized quasiparticle (soliton) that persists at zero temperature. The presence of thermal noise, however, could destabilize the protein soliton and dissipate its energy within a finite lifetime. In this work, we have computationally solved the system of stochastic (...) differential equations that govern the quantum dynamics of protein solitons at physiological temperature, T=310 K, for either a single isolated α-helix spine of hydrogen bonded peptide groups or the full protein α-helix comprised of three parallel α-helix spines. The simulated stochastic dynamics revealed that although the thermal noise is detrimental for the single isolated α-helix spine, the cooperative action of three amide I exciton quanta in the full protein α-helix ensures soliton lifetime of over 30 ps, during which the amide I energy could be transported along the entire extent of an 18-nm-long α-helix. Thus, macromolecular protein complexes, which are built up of protein α-helices could harness soliton-assisted energy transport at physiological temperature. Because the hydrolysis of a single adenosine triphosphate molecule is able to initiate three amide I exciton quanta, it is feasible that multiquantal protein solitons subserve a variety of specialized physiological functions in living systems. (shrink)

The mitogenic action of cytokines such as epidermal growth factor (EGF)d̊ or platelet dericed growth factor (PDGF) involves the stimulation of a signal cascade controlled by a small G protein called Ras. Mutations of Ras can cause its constitutive activation and, as a consequence, bypass the regulation of cell growth by cytokines. Both growth factor‐induced and oncogenic activation of Ras involve the conversion of Ras from the GDP‐bound (D‐Ras) to the GTP‐bound (T‐Ras) forms. T‐Ras activates a network of protein kinases (...) including c‐Mos, c‐Raf‐1 and MAP kinase. Eventually the activation of MAP kinase leads to the activation of the elongation factor 4E and several transcription factors such as c‐Jun, c‐Myc and c‐Fos. There are several modulators of Ras activity, such as the GTPase activating proteins (GAP1 and NF1), which stimulate the coversion of T‐Ras to D‐Ras. A series of small NF1 fragments, which bind T‐Ras, as well as truncated forms or derivatives of c‐Raf‐1, c‐Jun and c‐Myc, are capable of blocking the T‐Ras‐activated mitogenesis in a competitive manner. These agents offer a unique opportunity to control the proliferation of T‐Ras‐associated tumors, which represent more than 30% of total human carcinomas. (shrink)

The potential of the folic acid biosynthesis pathway as a target for the development of antibiotics has been acknowledged for many years and validated by the clinical use of several drugs. Recently, the crystal structures of all but one of the enzymes in the pathway from GTP to dihydrofolate have been determined. Given that structure‐based drug design strategies are now widely employed, these recent developments have prompted a re‐evaluation of the potential of each of the enzymes in the pathway as (...) a target for development of specific inhibitors. Here, we review the current knowledge of the structure and mechanism of each enzyme in the bacterial folic acid biosynthesis pathway from GTP to dihydrofolate and draw conclusions regarding the potential of each enzyme as a target for therapeutic intervention. BioEssays 24:637–648, 2002. © 2002 Wiley Periodicals, Inc. (shrink)

Phage DNA packaging occurs by DNA translocation into a prohead. Terminases are enzymes which initiate DNA packaging by cutting the DNA concatemer, and they are closely fitted structurally to the portal vertex of the prohead to form a ‘packasome’. Analysis among a number of phages supports an active role of the terminases in coupling ATP hydrolysis to DNA translocation through the portal. In phage T4 the small terminase subunit promotes a sequence‐specific terminase gene amplification within the chromosome. This link (...) between recombination and packaging suggests a DNA synapsis mechanism by the terminase to control packaging initiation, formally homologous to eukaryotic chromosome segregation. (shrink)

Transcription initiation by σ54–RNA polymerase (RNAP) relies explicitly on a transient interaction with a complex molecular machine belonging to the AAA+ (ATPases associated with various cellular activities) superfamily. Members of the AAA+ superfamily convert chemical energy derived from NTP hydrolysis to a mechanical force used to remodel their target substrate. Recently Bordes and colleagues,1 using a protein fragmentation approach, identified a unique sequence within σ54‐dependent transcriptional activators that constitutes a σ54‐binding interface. This interface is not static, but subject to (...) nucleotide‐dependent movement which may represent a common mechanism for controlling output that has been adopted by other AAA+ proteins. BioEssays 25:1150–1153, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

Phosphoinositides are inositol‐containing phospholipids whose hydrolysis is a key step in the rapid responses of animal cells to extracellular signals. Whether they play similar roles in plant cells has not been established, and some have suggested alternative roles as direct modulators of specific proteins. Nonetheless, evidence is accumulating that phosphoinositide hydrolysis mediates transduction of some signal in plants. The evidence is strongest for a role in triggering the shedding of flagella by the unicellular alga Chlamydomonas reinhardtii under acid (...) stress. Rapid kinetic analysis indicates that phosphoinositide hydrolysis occures within half a second and could trigger the rapid loss of flagella. Plant responses to pathogens and osmotic stress, as well as the regulation of turgor changes which underlie stomatal opening and closing and the movement of leaves and flower parts, may also be mediated by phosphoinositide hydrolysis. The evidence thus indicates that at least one reason plants have phosphoinositides is to mediate transduction of environmental signals. (shrink)

Proteins with seven conserved “helicase domains” play essential roles in all aspects of nucleic acid metabolism. Deriving energy from ATP hydrolysis, helicases alter the structure of DNA, RNA, or DNA:RNA duplexes, remodeling chromatin and modulating access to the DNA template by the transcriptional machinery. This review focuses on the diverse functions of these proteins in the process of RNA polymerase II transcription in eukaryotes. Known or putative helicases are required for general transcription initiation and for transcription-coupled DNA repair, and (...) may play important roles in elongation, termination, and transcript stability. Recent evidence suggests that helicase-domain-containing proteins are also involved in complexes that facilitate the activity of groups of seemingly unrelated genes. BioEssays 20:634–641, 1998. © 1998 John Wiley & Sons Inc. (shrink)

Completely updated and revised, the third edition of Catholic Health Care Ethics: A Manual for Practitioners sets the standard for Catholic bioethicists, physicians, nurses, and other health care workers. In thirty-nine chapters (many with subchapters), leading authors in their fields discuss a wide range of topics relevant to medicine and health care. The book has six parts covering foundational principles, health care ethics services, beginning-of-life issues, end-of-life issues, selected clinical issues, and institutional issues. Some highlights from the third edition include (...) new entries on the Ethical and Religious Directives for Catholic Health Care Services, certitude in moral decision-making, the principle of double effect, clinical ethics consultation, natural family planning, prenatal testing and diagnosis, care of fetal remains, challenges to neurological criteria, the use of ventilators, POLST, alkaline hydrolysis, opportunistic salpingectomy, so-called lethal prenatal diagnoses, transgenderism, and new age medicine. The volume continues to provide insightful information on the topics previously covered in the second edition, but with significant updates throughout. (shrink)

Understanding how chemical energy is converted into directed movement is a fundamental problem in biology. In higher organisms this is accomplished through the hydrolysis of ATP by three families of motor proteins: myosin, dynein and kinesin. The most abundant of these is myosin, which operates against actin and plays a central role in muscle contraction. As summarized here, great progress has been made towards understanding the molecular basis of movement through the determination of the three‐dimensional structures of myosin and (...) actin and through the establishment of systems for site‐directed mutagenesis of this motor protein. It now appears that the generation of movement is coupled to ATP hydrolysis by a series of domain movements within myosin. (shrink)

A large number of cellular proteins bind ATP, frequently utilizing the free energy of ATP hydrolysis to drive specific biological reactions. Recently, a family of closely related ATP‐binding proteins has been identified, the members of which share considerable sequence identity. These proteins, from both prokaryotic and eukaryotic sources, presumably had a common evolutionary origin and include the product of the white locus of Drosophila, the P‐glycoprotein which confers multidrug resistance on mammalian tumours, and prokaryotic proteins associated with such diverse (...) processes as membrane transport, cell division, nodulation and DNA repair. A comparison of these various proteins provides valuable insights into the function and evolution of the multicomponent systems with which they are associated. (shrink)

This commentary discusses the balance of phosphodiesterase and guanylate cyclase activities in vertebrate photoreceptors at moderate light intensities. The rate of cGMP hydrolysis and synthesis seem to equal each other. Ca2+as regulator of both enzyme activities is also effectively buffered in photoreceptor cells by cytoplasmic buffer components.

Segregation Distorter (SD) is a meiotic drive system in Drosophila that causes preferential transmission of the SD chromosome from SD/SD+ males owing to induced dysfunction of SD+ spermatids. Since its discovery in 1956, SD and its mode of action have baffled biologists. Recently, substantial progress has been made in elucidating this puzzle. Sd, the primary gene responsible for distortion encodes a mutant RanGAP, a key protein in the Ran signaling pathway required for nuclear transport and other nuclear functions. The mutant (...) protein is enzymatically active but mislocalized to nuclei, which apparently disrupts Ran signaling by reducing intranuclear Ran‐GTP levels. Some evidence suggests that a defect in nuclear transport may be the main cause of sperm dysfunction. Although important questions remain, the basic mechanism of distortion is now understood sufficiently well that specific hypotheses can be formulated and tested. This previously mysterious genetic system may now offer unique insights into novel aspects of regulation by Ran. BioEssays 25:108–115, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

Spontaneous polarization without spatial cues, or symmetry breaking, is a fundamental problem of spatial organization in biological systems. This question has been extensively studied using yeast models, which revealed the central role of the small GTPase switch Cdc42. Active Cdc42‐GTP forms a coherent patch at the cell cortex, thought to result from amplification of a small initial stochastic inhomogeneity through positive feedback mechanisms, which induces cell polarization. Here, I review and discuss the mechanisms of Cdc42 activity self‐amplification and dynamic turnover. (...) A robust Cdc42 patch is formed through the combined effects of Cdc42 activity promoting its own activation and active Cdc42‐GTP displaying reduced membrane detachment and lateral diffusion compared to inactive Cdc42‐GDP. I argue the role of the actin cytoskeleton in symmetry breaking is not primarily to transport Cdc42 to the active site. Finally, negative feedback and competition mechanisms serve to control the number of polarization sites. (shrink)

Multisubunit protein complexes are essential for cellular function. Genetic analysis of essential processes requires special tools, among which temperature‐sensitive (Ts) mutants have historically been crucial. Many researchers assume that the effect of temperature on such mutants is to drive their proteolytic destruction. In fact, degradation‐mediated elimination of mutant proteins likely explains only a fraction of the phenotypes associated with Ts mutants. Here I discuss insights gained from analysis of Ts mutants in oligomeric proteins, with particular focus on the study of (...) septins, GTP‐binding subunits of cytoskeletal filaments whose structures and functions are the subject of current investigation in my and many other labs. I argue that the kinds of unbiased forward genetic approaches that generate Ts mutants provide information that is largely inaccessible to modern reverse genetic methodologies, and will continue to drive our understanding of higher‐order assembly by septins and other oligomeric proteins.Also watch the Video Abstract. (shrink)

Base pair mismatches in DNA arise from errors in DNA replication, recombination, and biochemical modification of bases. Mismatches are inherently transient. They are resolved passively by DNA replication, or actively by enzymatic removal and resynthesis of one of the bases. The first step in removal is recognition of strand discontinuity by one of the MutS proteins. Mismatches arising from errors in DNA replication are repaired in favor of the base on the template strand, but other mismatches trigger base excision or (...) nucleotide excision repair (NER), or non‐repair pathways such as hypermutation, cell cycle arrest, or apoptosis. We argue that MutL homologues play a key role in determining biologic outcome by recruiting and/or activating effector proteins in response to lesion recognition by MutS. We suggest that the process is regulated by conformational changes in MutL caused by cycles of ATP binding and hydrolysis, and by physiologic changes which influence effector availability. (shrink)

The prenylated proteins represent a newly discovered class of post‐translationally modified proteins. The known prenylated proteins include the oncogene product p21ras and other low molecular weight GTP‐binding proteins, the nuclear lamins, and the γ subunit of the heterotrimeric G proteins. The modification involves the covalent attachment of a 15‐carbon (farnesyl) or 20‐carbon (geranylgeranyl) isoprenoid moiety in a thioether linkage to a carboxyl terminal cysteine. The nature of the attached substituent is dependent on specific sequence information in the carboxyl terminus of (...) the protein. In addition, prenylation entrains other post‐translational modifications forming a reaction pathway. In this article, we review our current understanding of the biochemical reactions involved in prenylation and discuss the possible role of this modification in the control of cellular functions such as protein maturation and cell growth. (shrink)

Neuronal growth and remodelling are guided by both intracellular gene programs and extracellular stimuli. The growth cone is one site where the effects of these extrinsic and intrinsic factors converge upon the mechanical determinants of cell shape. We review the growth cone as a transduction device, converting extracellular signals into mechanical forces. A variety of soluble, extracellular matrix and membrane bound molecules control growth cone behavior. In addition, GAP‐43 is discussed as a possible component of the Intraneuronal gene program which (...) modulates growth cone activity. The GTP‐binding protein, Go, is a major growth cone membrane protein that may transduce signals not only from outside the cell, but from within as well. This may provide a molecular site in the growth cone for the coordination of a genetic growth program with environmental signals. (shrink)

In the current model of visual transduction, the lifetime of active cGMP phosphodiesterase depends upon the period of its interaction with GTP-bound transducin. If recoverin regulates the lifetime of light-activated cGMP phosphodiesterase through inhibition of rhodopsin phosphorylation, rhodopsin should directly interact with cGMP phosphodiesterase and/or GTP-bound transducin complexed with cGMP phosphodiesterase. Is this true?

Sarco/endoplasmic reticulum Ca2+ ATPase 2b (SERCA2b), a member of the SERCA family, is expressed ubiquitously and transports Ca2+ into the sarco/endoplasmic reticulum using the energy provided by ATP binding and hydrolysis. The crystal structure of SERCA2b in its Ca2+‐ and ATP‐bound (E1∙2Ca2+‐ATP) state and cryo‐electron microscopy (cryo‐EM) structures of the protein in its E1∙2Ca2+‐ATP and Ca2+‐unbound phosphorylated (E2P) states have provided essential insights into how the overall conformation and ATPase activity of SERCA2b is regulated by the transmembrane helix 11 (...) and the subsequent luminal extension loop, both of which are specific to this isoform. More recently, our cryo‐EM analysis has revealed that SERCA2b likely adopts open and closed conformations of the cytosolic domains in the Ca2+‐bound but ATP‐free (E1∙2Ca2+) state, and that the closed conformation represents a state immediately prior to ATP binding. This review article summarizes the unique mechanisms underlying the conformational and functional regulation of SERCA2b. (shrink)

Currently, a central question in biology is how signals from the cell surface modulate intracellular processes. In recent years phosphoinositides have been shown to play a key role in signal transduction. Two phosphoinositide pathways have been characterized, to date. In the canonical phosphoinositide turnover pathway, activation of phosphatidylinositol‐specific phospholipase C results in the hydrolysis of phosphatidylinositol 4,5‐bisphospate and the generation of two second messengers, inositol 1,4,5‐trisphosphate and diacylglycerol. The 3‐phosphoinositide pathway involves protein‐tyrosine kinase‐mediated recruitment and activation of phosphatidylinositol 3‐kinase, (...) resulting in the production of phosphatidylinositol 3,4‐bisphosphate and phosphatidylinositol 3,4,5‐trisphosphate. The 3‐phosphoinositides are not substrates of any known phospholipase C, are not components of the canonical phosphoinositide turnover pathway, and may themselves act as intracellular mediators. The 3‐phosphoinositide pathway has been implicated in growth factor‐dependent mitogenesis, membrane ruffling and glucose uptake. Furthermore the homology of the yeast vps34 with the mammalian phosphatidylinositol 3‐kinase has suggested a role for this pathway in vesicular trafficking.In this review the different mechanisms employed by protein‐tyrosine kinases to activate phosphatidylinositol 3‐kinase, and its involvement in the signaling cascade initiated by tyrosine phosphorylation, are examined. (shrink)

It was, until recently, accepted that the two classes of acetylcholine (ACh) receptors are distinct in an important sense: muscarinic ACh receptors signal via heterotrimeric GTP binding proteins (G proteins), whereas nicotinic ACh receptors (nAChRs) open to allow flux of Na+, Ca2+, and K+ ions into the cell after activation. Here we present evidence of direct coupling between G proteins and nAChRs in neurons. Based on proteomic, biophysical, and functional evidence, we hypothesize that binding to G proteins modulates the activity (...) and signaling of nAChRs in cells. It is important to note that while this hypothesis is new for the nAChR, it is consistent with known interactions between G proteins and structurally related ligand‐gated ion channels. Therefore, it underscores an evolutionarily conserved metabotropic mechanism of G protein signaling via nAChR channels.Also watch the Video Abstract. (shrink)

It was, until recently, accepted that the two classes of acetylcholine (ACh) receptors are distinct in an important sense: muscarinic ACh receptors signal via heterotrimeric GTP binding proteins (G proteins), whereas nicotinic ACh receptors (nAChRs) open to allow flux of Na+, Ca2+, and K+ ions into the cell after activation. Here we present evidence of direct coupling between G proteins and nAChRs in neurons. Based on proteomic, biophysical, and functional evidence, we hypothesize that binding to G proteins modulates the activity (...) and signaling of nAChRs in cells. It is important to note that while this hypothesis is new for the nAChR, it is consistent with known interactions between G proteins and structurally related ligand‐gated ion channels. Therefore, it underscores an evolutionarily conserved metabotropic mechanism of G protein signaling via nAChR channels.Also watch the Video Abstract. (shrink)

The Rho GTPases—Rho, Rac and Cdc42—act as molecular switches, cycling between an active GTP‐bound state and an inactive GDP‐bound state, to regulate the actin cytoskeleton. It has recently become apparent that the Rho GTPases can be activated in subcellular zones that appear semi‐stable, yet are dynamically maintained. These Rho GTPase activity zones are associated with a variety of fundamental biological processes including symmetric and asymmetric cytokinesis and cellular wound repair. Here we review the basic features of Rho GTPase activity zones, (...) suggest that these zones represent a fundamental signaling mechanism, and discuss the implications of zone properties from the perspective of both their function and how they are likely to be controlled. BioEssays 28: 983–993, 2006. © 2006 Wiley Periodicals, Inc. (shrink)

Light activated rhodopsin functions by catalyzing the exchange of GTP for GDP on numerous copies of transducin. Peptide mapping has shown that at least six regions, three on rhodopsin and three on the transducin alpha subunit, are involved in the active interface between the two proteins. The most informative structural studies of rhodopsin should include focus on the transducin interaction.

Molecular motor proteins, fueled by energy from ATP hydrolysis, move along actin filaments or microtubules, performing work in the cell. The kinesin microtubule motors transport vesicles or organelles, assemble bipolar spindles or depolymerize microtubules, functioning in basic cellular processes. The mechanism by which motor proteins convert energy from ATP hydrolysis into work is likely to differ in basic ways from man‐made machines. Several mechanical elements of the kinesin motors have now been tentatively identified, permitting researchers to begin to (...) decipher the mechanism of motor function. The force‐producing conformational changes of the motor and the means by which they are amplified are probably different for the plus‐ and minus‐end kinesin motors. BioEssays 25:1212–1219, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

The cellular machinery responsible for conveying proteins between the endoplasmic reticulum and the Golgi is being investigated using genetics and biochemistry. A role for vesicles in mediating protein traffic between the ER and the Golgi has been established by characterizing yeast mutants defective in this process, and by using recently developed cell‐free assays that measure ER to Golgi transport. These tools have also allowed the identification of several proteins crucial to intracellular protein trafficking. The characterization and possible functions of several (...) GTP‐binding proteins, peripheral membrane proteins, and an integral membrane protein during ER to Golgi transport are discussed here. (shrink)