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The Malpighian tubules of Calliphora are described, emphasizing the possible role of surface specializations in solute-linked water transport. The tubules are composed of two cell types, primary and stellate, intermingling along the tubule length. The primary cells have long narrow basal infoldings and a microvillate luminal border, both intimately associated with mitochondria. The stellate cells have shorter and wider basal infoldings and their apical microvilli do not contain mitochondria. Application of the standing gradient hypothesis to this sytem provides a model for urine formation in which the local gradients for osmotic water flow occur within the long narrow channels of the basal infolds and microvilli of the primary cells. Stellate cells may modify the initial secretion by reabsorbing sodium. 相似文献
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Holmes KC Geeves MA 《Philosophical transactions of the Royal Society of London. Series B, Biological sciences》2000,355(1396):419-431
The myosin cross-bridge exists in two conformations, which differ in the orientation of a long lever arm. Since the lever arm undergoes a 60 degree rotation between the two conformations, which would lead to a displacement of the myosin filament of about 11 nm, the transition between these two states has been associated with the elementary 'power stroke' of muscle. Moreover, this rotation is coupled with changes in the active site (CLOSED to OPEN), which probably enable phosphate release. The transition CLOSED to OPEN appears to be brought about by actin binding. However, kinetics shows that the binding of myosin to actin is a two-step process which affects both ATP and ADP affinity and vice versa. The structural basis of these effects is only partially explained by the presently known conformers of myosin. Therefore, additional states of the myosin cross-bridge should exist. Indeed, cryoelectron microscopy has revealed other angles of the lever arm induced by ADP binding to a smooth muscle actin-myosin complex. 相似文献
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The structural basis of molecular adaptation 总被引:31,自引:21,他引:10
The study of molecular adaptation has long been fraught with difficulties,
not the least of which is identifying out of hundreds of amino acid
replacements those few directly responsible for major adaptations. Six
studies are used to illustrate how phylogenies, site- directed mutagenesis,
and a knowledge of protein structure combine to provide much deeper
insights into the adaptive process than has hitherto been possible. Ancient
genes can be reconstructed, and the phenotypes can be compared to modern
proteins. Out of hundreds of amino acid replacements accumulated over
billions of years those few responsible for discriminating between
alternative substrates are identified. An amino acid replacement of modest
effect at the molecular level causes a dramatic expansion in an ecological
niche. These and other topics are creating the emerging field of
"paleomolecular biochemistry."
相似文献
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To deliver their genetic material into host cells, enveloped viruses have surface glycoproteins that actively cause the fusion of the viral and cellular membranes. Recently determined X-ray crystal structures of the paramyxovirus fusion (F) protein in its pre-fusion and post-fusion conformations reveal the dramatic structural transformation that this protein undergoes while causing membrane fusion. Conformational changes in key regions of the F protein suggest the mechanism by which the F protein is activated and refolds. 相似文献
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Sirtuins comprise a family of enzymes that catalyze the deacetylation of acetyllysine side chains in a reaction that consumes NAD+. Although several crystal structures of sirtuins bound to non-native acetyl peptides have been determined, relatively little about how sirtuins discriminate among different substrates is understood. We have carried out a systematic structural and thermodynamic analysis of several peptides bound to a single sirtuin, the Sir2 homologue from Thermatoga maritima (Sir2Tm). We report structures of five different forms of Sir2Tm: two forms bound to the p53 C-terminal tail in the acetylated and unacetylated states, two forms bound to putative acetyl peptide substrates derived from the structured domains of histones H3 and H4, and one form bound to polypropylene glycol (PPG), which resembles the apoenzyme. The structures reveal previously unobserved complementary side chain interactions between Sir2Tm and the first residue N-terminal to the acetyllysine (position -1) and the second residue C-terminal to the acetyllysine (position +2). Isothermal titration calorimetry was used to compare binding constants between wild-type and mutant forms of Sir2Tm and between additional acetyl peptide substrates with substitutions at positions -1 and +2. The results are consistent with a model in which peptide positions -1 and +2 play a significant role in sirtuin substrate binding. This model provides a framework for identifying sirtuin substrates. 相似文献
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Srivastava D Singh RK Moxley MA Henzl MT Becker DF Tanner JJ 《Journal of molecular biology》2012,420(3):176-189
Type II hyperprolinemia is an autosomal recessive disorder caused by a deficiency in Δ1-pyrroline-5-carboxylate dehydrogenase (P5CDH; also known as ALDH4A1), the aldehyde dehydrogenase that catalyzes the oxidation of glutamate semialdehyde to glutamate. Here, we report the first structure of human P5CDH (HsP5CDH) and investigate the impact of the hyperprolinemia-associated mutation of Ser352 to Leu on the structure and catalytic properties of the enzyme. The 2. 5-Å-resolution crystal structure of HsP5CDH was determined using experimental phasing. Structures of the mutant enzymes S352A (2.4 Å) and S352L (2.85 Å) were determined to elucidate the structural consequences of altering Ser352. Structures of the 93% identical mouse P5CDH complexed with sulfate ion (1.3 Å resolution), glutamate (1.5 Å), and NAD+ (1.5 Å) were determined to obtain high-resolution views of the active site. Together, the structures show that Ser352 occupies a hydrophilic pocket and is connected via water-mediated hydrogen bonds to catalytic Cys348. Mutation of Ser352 to Leu is shown to abolish catalytic activity and eliminate NAD+ binding. Analysis of the S352A mutant shows that these functional defects are caused by the introduction of the nonpolar Leu352 side chain rather than the removal of the Ser352 hydroxyl. The S352L structure shows that the mutation induces a dramatic 8-Å rearrangement of the catalytic loop. Because of this conformational change, Ser349 is not positioned to interact with the aldehyde substrate, conserved Glu447 is no longer poised to bind NAD+, and Cys348 faces the wrong direction for nucleophilic attack. These structural alterations render the enzyme inactive. 相似文献
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Secondary active transporters couple the free energy of the electrochemical potential of one solute to the transmembrane movement of another. As a basic mechanistic explanation for their transport function the model of alternating access was put forward more than 40 years ago, and has been supported by numerous kinetic, biochemical and biophysical studies. According to this model, the transporter exposes its substrate binding site(s) to one side of the membrane or the other during transport catalysis, requiring a substantial conformational change of the carrier protein. In the light of recent structural data for a number of secondary transport proteins, we analyze the model of alternating access in more detail, and correlate it with specific structural and chemical properties of the transporters, such as their assignment to different functional states in the catalytic cycle of the respective transporter, the definition of substrate binding sites, the type of movement of the central part of the carrier harboring the substrate binding site, as well as the impact of symmetry on fold-specific conformational changes. Besides mediating the transmembrane movement of solutes, the mechanism of secondary carriers inherently involves a mechanistic coupling of substrate flux to the electrochemical potential of co-substrate ions or solutes. Mainly because of limitations in resolution of available transporter structures, this important aspect of secondary transport cannot yet be substantiated by structural data to the same extent as the conformational change aspect. We summarize the concepts of coupling in secondary transport and discuss them in the context of the available evidence for ion binding to specific sites and the impact of the ions on the conformational state of the carrier protein, which together lead to mechanistic models for coupling. 相似文献
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The structural basis of light-harvesting in purple bacteria 总被引:6,自引:0,他引:6
Cogdell RJ Isaacs NW Freer AA Howard TD Gardiner AT Prince SM Papiz MZ 《FEBS letters》2003,555(1):35-39
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Allosteric regulation of protein function occurs when the regulatory trigger, such as the binding of a small-molecule effector or inhibitor, takes place some distance from the protein’s, or protein complex’s, active site. This distance can be a few Å, or tens of Å. Many proteins are regulated in this way and exhibit a variety of allosteric mechanisms. Here we review how analyses of experimentally determined models of protein 3D structures, using either X-ray crystallography or NMR spectroscopy, have revealed some of the mechanisms involved. 相似文献
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Marcia E. Newcomer Alan R. Brash 《Protein science : a publication of the Protein Society》2015,24(3):298-309
Many intriguing facets of lipoxygenase (LOX) catalysis are open to a detailed structural analysis. Polyunsaturated fatty acids with two to six double bonds are oxygenated precisely on a particular carbon, typically forming a single chiral fatty acid hydroperoxide product. Molecular oxygen is not bound or liganded during catalysis, yet it is directed precisely to one position and one stereo configuration on the reacting fatty acid. The transformations proceed upon exposure of substrate to enzyme in the presence of O2 (RH + O2 → ROOH), so it has proved challenging to capture the precise mode of substrate binding in the LOX active site. Beginning with crystal structures with bound inhibitors or surrogate substrates, and most recently arachidonic acid bound under anaerobic conditions, a picture is consolidating of catalysis in a U‐shaped fatty acid binding channel in which individual LOX enzymes use distinct amino acids to control the head‐to‐tail orientation of the fatty acid and register of the selected pentadiene opposite the non‐heme iron, suitably positioned for the initial stereoselective hydrogen abstraction and subsequent reaction with O2. Drawing on the crystal structures available currently, this review features the roles of the N‐terminal β‐barrel (C2‐like, or PLAT domain) in substrate acquisition and sensitivity to cellular calcium, and the α‐helical catalytic domain in fatty acid binding and reactions with O2 that produce hydroperoxide products with regio and stereospecificity. LOX structures combine to explain how similar enzymes with conserved catalytic machinery differ in product, but not substrate, specificities. 相似文献
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Robert H Brown David B Pearse George Pyrgos Mark C Liu Alkis Togias Solbert Permutt 《Journal of applied physiology》2006,101(1):30-39
We hypothesized that structural airway remodeling contributes to airways hyperresponsiveness (AHR) in asthma. Small, medium, and large airways were analyzed by computed tomography in 21 asthmatic volunteers under baseline conditions (FEV1 = 64% predicted) and after maximum response to albuterol (FEV1 = 76% predicted). The difference in pulmonary function between baseline and albuterol was an estimate of AHR to the baseline smooth muscle tone (BSMT). BSMT caused an increase in residual volume (RV) that was threefold greater than the decrease in forced vital capacity (FVC) because of a simultaneous increase in total lung capacity (TLC). The decrease in FVC with BSMT was the major determinant of the baseline FEV1 (P < 0.0001). The increase in RV correlated inversely with the relaxed luminal diameter of the medium airways (P = 0.009) and directly with the wall thickness of the large airways (P = 0.001). The effect of BSMT on functional residual capacity (FRC) controlled the change in TLC relative to the change in RV. When the FRC increased with RV, TLC increased and FVC was preserved. When the relaxed large airways were critically narrowed, FRC and TLC did not increase and FVC fell. With critical large airways narrowing, the FRC was already elevated from dynamic hyperinflation before BSMT and did not increase further with BSMT. FEV1/FVC in the absence of BSMT correlated directly with large airway luminal diameter and inversely with the fall in FVC with BSMT. These findings suggest that dynamic hyperinflation caused by narrowing of large airways is a major determinant of AHR in asthma. 相似文献
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The structural basis of the TIM10 chaperone assembly 总被引:5,自引:0,他引:5
Lu H Golovanov AP Alcock F Grossmann JG Allen S Lian LY Tokatlidis K 《The Journal of biological chemistry》2004,279(18):18959-18966
Tim9 and Tim10 are essential components of the "small Tim" family of proteins that facilitate insertion of polytopic proteins at the inner mitochondrial membrane. The small Tims are themselves imported from the cytosol and are organized in specific translocation assemblies in the intermembrane space. Their conformational properties and how these influence the mechanism of assembly remain poorly understood. Moreover, the three-dimensional structure of the TIM10 complex is unknown. We have characterized the structural properties of these proteins in their free and assembled states using NMR, circular dichroism, and small angle x-ray scattering. We show that the free proteins are largely unfolded in their reduced assembly-incompetent state and molten globules in their oxidized assembly-competent state. Tim10 appears less structured than Tim9 in their respective free oxidized forms and undergoes a larger structural change than Tim9 upon complexation. The NMR data here demonstrates unequivocally that only the oxidized states of the Tim9 and Tim10 proteins are capable of forming a complex. Zinc binding stabilizes the reduced state against proteolysis without significantly affecting the secondary structure. Solution x-ray scattering was used to obtain a molecular envelope for the subunits individually and for their fully functional TIM10 complex. Ab initio shape reconstructions based on the scattering data has allowed us to obtain the first low resolution three-dimensional structure of the TIM10 complex. This is a novel structure that displays extensive surface hydrophobicity. The structure also provides an explanation for the escorting function of this non-ATP-powered chaperone particle. 相似文献
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Photosynthesis Research - Phycobilisomes (PBS), the major light-harvesting antenna in cyanobacteria, are supramolecular complexes of colorless linkers and heterodimeric, pigment-binding... 相似文献