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1.
A tight coupling between adenosine triphosphate (ATP) hydrolysis and vectorial ion transport has to be maintained by ATP-consuming ion pumps. We report two crystal structures of Ca2+-bound sarco(endo)plasmic reticulum Ca2+-adenosine triphosphatase (SERCA) at 2.6 and 2.9 angstrom resolution in complex with (i) a nonhydrolyzable ATP analog [adenosine (beta-gamma methylene)-triphosphate] and (ii) adenosine diphosphate plus aluminum fluoride. SERCA reacts with ATP by an associative mechanism mediated by two Mg2+ ions to form an aspartyl-phosphorylated intermediate state (Ca2-E1 approximately P). The conformational changes that accompany the reaction with ATP pull the transmembrane helices 1 and 2 and close a cytosolic entrance for Ca2+, thereby preventing backflow before Ca2+ is released on the other side of the membrane. 相似文献
2.
Proton pumps in the plasma membrane of plants and yeasts maintain the intracellular pH and membrane potential. To gain insight into the molecular mechanisms of proton pumping, we built an atomic homology model of the proton pump based on the 2.6 angstrom x-ray structure of the related Ca2+ pump from rabbit sarcoplasmic reticulum. The model, when fitted to an 8 angstrom map of the Neurospora proton pump determined by electron microscopy, reveals the likely path of the proton through the membrane and shows that the nucleotide-binding domain rotates by approximately 70 degrees to deliver adenosine triphosphate (ATP) to the phosphorylation site. A synthetic peptide corresponding to the carboxyl-terminal regulatory domain stimulates ATPase activity, suggesting a mechanism for proton transport regulation. 相似文献
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4.
Sambongi Y Iko Y Tanabe M Omote H Iwamoto-Kihara A Ueda I Yanagida T Wada Y Futai M 《Science (New York, N.Y.)》1999,286(5445):1722-1724
F0F1, found in mitochondria or bacterial membranes, synthesizes adenosine 5'-triphosphate (ATP) coupling with an electrochemical proton gradient and also reversibly hydrolyzes ATP to form the gradient. An actin filament connected to a c subunit oligomer of F0 was able to rotate by using the energy of ATP hydrolysis. The rotary torque produced by the c subunit oligomer reached about 40 piconewton-nanometers, which is similar to that generated by the gamma subunit in the F1 motor. These results suggest that the gamma and c subunits rotate together during ATP hydrolysis and synthesis. Thus, coupled rotation may be essential for energy coupling between proton transport through F0 and ATP hydrolysis or synthesis in F1. 相似文献
5.
The dynamics and polarity of actin filaments are controlled by a conformational change coupled to the hydrolysis of adenosine 5'-triphosphate (ATP) by a mechanism that remains to be elucidated. Actin modified to block polymerization was crystallized in the adenosine 5'-diphosphate (ADP) state, and the structure was solved to 1.54 angstrom resolution. Compared with previous ATP-actin structures from complexes with deoxyribonuclease I, profilin, and gelsolin, monomeric ADP-actin is characterized by a marked conformational change in subdomain 2. The successful crystallization of monomeric actin opens the way to future structure determinations of actin complexes with actin-binding proteins such as myosin. 相似文献
6.
Adenosine triphosphate (ATP)-binding cassette (ABC) transporters convert chemical energy from ATP hydrolysis to mechanical work for substrate translocation. They function by alternating between two states, exposing the substrate-binding site to either side of the membrane. A key question that remains to be addressed is how substrates initiate the transport cycle. Using x-ray crystallography, we have captured the maltose transporter in an intermediate step between the inward- and outward-facing states. We show that interactions with substrate-loaded maltose-binding protein in the periplasm induce a partial closure of the MalK dimer in the cytoplasm. ATP binding to this conformation then promotes progression to the outward-facing state. These results, interpreted in light of biochemical and functional studies, provide a structural basis to understand allosteric communication in ABC transporters. 相似文献
7.
The crystal structure of the high-affinity Escherichia coli MetNI methionine uptake transporter, a member of the adenosine triphosphate (ATP)-binding cassette (ABC) family, has been solved to 3.7 angstrom resolution. The overall architecture of MetNI reveals two copies of the adenosine triphosphatase (ATPase) MetN in complex with two copies of the transmembrane domain MetI, with the transporter adopting an inward-facing conformation exhibiting widely separated nucleotide binding domains. Each MetI subunit is organized around a core of five transmembrane helices that correspond to a subset of the helices observed in the larger membrane-spanning subunits of the molybdate (ModBC) and maltose (MalFGK) ABC transporters. In addition to the conserved nucleotide binding domain of the ABC family, MetN contains a carboxyl-terminal extension with a ferredoxin-like fold previously assigned to a conserved family of regulatory ligand-binding domains. These domains separate the nucleotide binding domains and would interfere with their association required for ATP binding and hydrolysis. Methionine binds to the dimerized carboxyl-terminal domain and is shown to inhibit ATPase activity. These observations are consistent with an allosteric regulatory mechanism operating at the level of transport activity, where increased intracellular levels of the transported ligand stabilize an inward-facing, ATPase-inactive state of MetNI to inhibit further ligand translocation into the cell. 相似文献
8.
Sano Y Inamura K Miyake A Mochizuki S Yokoi H Matsushime H Furuichi K 《Science (New York, N.Y.)》2001,293(5533):1327-1330
We characterized an activation mechanism of the human LTRPC2 protein, a member of the transient receptor potential family of ion channels, and demonstrated that LTRPC2 mediates Ca2+ influx into immunocytes. Intracellular pyrimidine nucleotides, adenosine 5'-diphosphoribose (ADPR), and nicotinamide adenine dinucleotide (NAD), directly activated LTRPC2, which functioned as a Ca2+-permeable nonselective cation channel and enabled Ca2+ influx into cells. This activation was suppressed by intracellular adenosine triphosphate. These results reveal that ADPR and NAD act as intracellular messengers and may have an important role in Ca2+ influx by activating LTRPC2 in immunocytes. 相似文献
9.
Protein kinase activity closely associated with a reconstituted calcium-activated potassium channel. 总被引：14，自引：0，他引：14
S K Chung P H Reinhart B L Martin D Brautigan I B Levitan 《Science (New York, N.Y.)》1991,253(5019):560-562
Modulation of the activity of potassium and other ion channels is an essential feature of nervous system function. The open probability of a large conductance Ca(2+)-activated K+ channel from rat brain, incorporated into planar lipid bilayers, is increased by the addition of adenosine triphosphate (ATP) to the cytoplasmic side of the channel. This modulation takes place without the addition of protein kinase, requires Mg2+, and is mimicked by an ATP analog that serves as a substrate for protein kinases but not by a nonhydrolyzable ATP analog. Addition of protein phosphatase 1 reverses the modulation by MgATP. Thus, there may be an endogenous protein kinase activity firmly associated with this K+ channel. Some ion channels may exist in a complex that contains regulatory protein kinases and phosphatases. 相似文献
10.
Select members of the adenosine triphosphate (ATP)-binding cassette (ABC) transporter family couple ATP binding and hydrolysis to substrate efflux and confer multidrug resistance. We have determined the x-ray structure of MsbA in complex with magnesium, adenosine diphosphate, and inorganic vanadate (Mg.ADP.Vi) and the rough-chemotype lipopolysaccharide, Ra LPS. The structure supports a model involving a rigid-body torque of the two transmembrane domains during ATP hydrolysis and suggests a mechanism by which the nucleotide-binding domain communicates with the transmembrane domain. We propose a lipid "flip-flop" mechanism in which the sugar groups are sequestered in the chamber while the hydrophobic tails are dragged through the lipid bilayer. 相似文献
11.
The motor protein kinesin moves along microtubules, driven by adenosine triphosphate (ATP) hydrolysis. However, it remains unclear how kinesin converts the chemical energy into mechanical movement. We report crystal structures of monomeric kinesin KIF1A with three transition-state analogs: adenylyl imidodiphosphate (AMP-PNP), adenosine diphosphate (ADP)-vanadate, and ADP-AlFx (aluminofluoride complexes). These structures, together with known structures of the ADP-bound state and the adenylyl-(beta,gamma-methylene) diphosphate (AMP-PCP)-bound state, show that kinesin uses two microtubule-binding loops in an alternating manner to change its interaction with microtubules during the ATP hydrolysis cycle; loop L11 is extended in the AMP-PNP structure, whereas loop L12 is extended in the ADP structure. ADP-vanadate displays an intermediate structure in which a conformational change in two switch regions causes both loops to be raised from the microtubule, thus actively detaching kinesin. 相似文献
12.
p-nitrophenyl phosphate hydrolysis and calcium ion transport in fragmented sarcoplasmic reticulum 总被引：3，自引：0，他引：3
G Inesi 《Science (New York, N.Y.)》1971,171(974):901-903
The calcium ion pump of fragmented sarcoplasmic reticulum can be coupled to hydrolysis of p-nitrophenyl phosphate, in the absence of added adenosine triphosphate. Comparison of the activities obtained with the two substrates suggests an analogous mechanism of transport. Independent of the substrate, a 2 : 1 ratio between calcium ion transport and substrate hydrolysis is displayed by the system, and an identical amount of work is required for ion transport against a given gradient. A phosphate ester appears necessary for substrate utilization in the pump mechanism, whereas the structure of the substrate determines the rates of activity and the affinity of the system for calcium ion. 相似文献
13.
Oxygen consumption and cellular ion transport: evidence for adenosine triphosphate to O2 ratio near 6 in intact cell 总被引：4，自引：0，他引：4
Oxygen (O2) consumption and net K+ uptake were measured simultaneously upon reintroduction of K+ into a K+-depleted suspension of renal tubules. The K+/O2 stoichiometries of 11.8 +/- 0.2 and 8.4 +/- 0.6 were obtained for reduced nicotinamide adenine dinucleotide- and flavoprotein-linked substrates, respectively. These values complement classical K+ to adenosine triphosphate (ATP) and ATP/O2 stoichiometries, thereby demonstrating a remarkably efficient coupling between the processes of Na+- and K+-dependent adenosinetriphosphatase-mediated ion transport and oxidative phosphorylation within the intact cell. 相似文献
14.
Processive chromosomal replication relies on sliding DNA clamps, which are loaded onto DNA by pentameric clamp loader complexes belonging to the AAA+ family of adenosine triphosphatases (ATPases). We present structures for the ATP-bound state of the clamp loader complex from bacteriophage T4, bound to an open clamp and primer-template DNA. The clamp loader traps a spiral conformation of the open clamp so that both the loader and the clamp match the helical symmetry of DNA. One structure reveals that ATP has been hydrolyzed in one subunit and suggests that clamp closure and ejection of the loader involves disruption of the ATP-dependent match in symmetry. The structures explain how synergy among the loader, the clamp, and DNA can trigger ATP hydrolysis and release of the closed clamp on DNA. 相似文献
15.
Actin polymerization and ATP hydrolysis 总被引：18，自引：0，他引：18
F-actin is the major component of muscle thin filaments and, more generally, of the microfilaments of the dynamic, multifunctional cytoskeletal systems of nonmuscle eukaryotic cells. Polymeric F-actin is formed by reversible noncovalent self-association of monomeric G-actin. To understand the dynamics of microfilament systems in cells, the dynamics of polymerization of pure actin must be understood. The following model has emerged from recent work. During the polymerization process, adenosine 5'-triphosphate (ATP) that is bound to G-actin is hydrolyzed to adenosine 5'-diphosphate (ADP) that is bound to F-actin. The hydrolysis reaction occurs on the F-actin subsequent to the polymerization reaction in two steps: cleavage of ATP followed by the slower release of inorganic phosphate (Pi). As a result, at high rates of filament growth a transient cap of ATP-actin subunits exists at the ends of elongating filaments, and at steady state a stabilizing cap of ADP.Pi-actin subunits exists at the barbed ends of filaments. Cleavage of ATP results in a highly stable filament with bound ADP.Pi, and release of Pi destabilizes the filament. Thus these two steps of the hydrolytic reaction provide potential mechanisms for regulating the monomer-polymer transition. 相似文献
16.
The motility of kinesin motors is explained by a "hand-over-hand" model in which two heads of kinesin alternately repeat single-headed and double-headed binding with a microtubule. To investigate the binding mode of kinesin at the key nucleotide states during adenosine 5'-triphosphate (ATP) hydrolysis, we measured the mechanical properties of a single kinesin-microtubule complex by applying an external load with optical tweezers. Both the unbinding force and the elastic modulus in solutions containing AMP-PNP (an ATP analog) were twice the value of those in nucleotide-free solution or in the presence of both AMP-PNP and adenosine 5'-diphosphate. Thus, kinesin binds through two heads in the former and one head in the latter two states, which supports a major prediction of the hand-over-hand model. 相似文献
17.
D L Alkon J Acosta-Urquidi J Olds G Kuzma J T Neary 《Science (New York, N.Y.)》1983,219(4582):303-306
Intracellular iontophoretic injection of the catalytic subunit of cyclic adenosine monophosphate-dependent protein kinase increased input resistance and decreased a delayed voltage-dependent K+ current of the type B photoreceptor in the nudibranch Hermissenda crassicornis to a greater extent than an early, rapidly inactivating K+ current (IA). This injection also enhanced the long-lasting depolarization of type B cells after a light step. These findings suggest the involvement of cyclic adenosine monophosphate-dependent phosphorylation in the differential regulation of photoreceptor K+ currents particularly during illumination. On the other hand, conditioning-induced changes in IA may also be regulated by a different type of phosphorylation (for example, Ca2+-dependent). 相似文献
18.
Tezcan FA Kaiser JT Mustafi D Walton MY Howard JB Rees DC 《Science (New York, N.Y.)》2005,309(5739):1377-1380
Adenosine triphosphate (ATP) hydrolysis in the nitrogenase complex controls the cycle of association and dissociation between the electron donor adenosine triphosphatase (ATPase) (Fe-protein) and its target catalytic protein (MoFe-protein), driving the reduction of dinitrogen into ammonia. Crystal structures in different nucleotide states have been determined that identify conformational changes in the nitrogenase complex during ATP turnover. These structures reveal distinct and mutually exclusive interaction sites on the MoFe-protein surface that are selectively populated, depending on the Fe-protein nucleotide state. A consequence of these different docking geometries is that the distance between redox cofactors, a critical determinant of the intermolecular electron transfer rate, is coupled to the nucleotide state. More generally, stabilization of distinct docking geometries by different nucleotide states, as seen for nitrogenase, could enable nucleotide hydrolysis to drive the relative motion of protein partners in molecular motors and other systems. 相似文献
19.
Studies and perspectives of protein kinase C 总被引：335，自引：0，他引：335
Y Nishizuka 《Science (New York, N.Y.)》1986,233(4761):305-312
Protein kinase C, an enzyme that is activated by the receptor-mediated hydrolysis of inositol phospholipids, relays information in the form of a variety of extracellular signals across the membrane to regulate many Ca2+-dependent processes. At an early phase of cellular responses, the enzyme appears to have a dual effect, providing positive forward as well as negative feedback controls over various steps of its own and other signaling pathways, such as the receptors that are coupled to inositol phospholipid hydrolysis and those of some growth factors. In biological systems, a positive signal is frequently followed by immediate negative feedback regulation. Such a novel role of this protein kinase system seems to give a logical basis for clarifying the biochemical mechanism of signal transduction, and to add a new dimension essential to our understanding of cell-to-cell communication. 相似文献
20.
DNA gyrase and the supercoiling of DNA 总被引：101，自引：0，他引：101
N R Cozzarelli 《Science (New York, N.Y.)》1980,207(4434):953-960
Negative supercoiling of bacterial DNA by DNA gyrase influences all metabolic processes involving DNA and is essential for replication. Gyrase supercoils DNA by a mechanism called sign inversion, whereby a positive supercoil is directly inverted to a negative one by passing a DNA segment through a transient double-strand break. Reversal of this scheme relaxes DNA, and this mechanism also accounts for the ability of gyrase to catenate and uncatenate DNA rings. Each round of supercoiling is driven by a conformational change induced by adenosine triphosphate (ATP) binding: ATP hydrolysis permits fresh cycles. The inhibition of gyrase by two classes of antimicrobials reflects its composition from two reversibly associated subunits. The A subunit is particularly associated with the concerted breakage-and-rejoining of DNA and the B subunit mediates energy transduction. Gyrase is a prototype for a growing class of prokaryotic and eukaryotic topoisomerases that interconvert complex forms by way of transient double-strand breaks. 相似文献