The influence of salt concentration on CD spectra of tRNA and aminoacyl-tRNA synthetase

The CD spectra of serine tRNA or seryl-tRNA synthetase were measured. The [theta] values at 210 nm were minimum at 50mM - 0.2M NaCl, at that concentration the velocity of aminoacylation was maximum. This results suggest that A . U and G . C base pairs loosened. The [theta] values at 200 nm decreased according to the decreasing of salt concentration, suggesting the decomposition of A . U base pairs. The CD spectra of seryl-tRNA synthetase at 210-240 nm were not changed in the range of 10mM-0.3M NaCl but the spectra at 260-290 nm showed minimum in the range between 50mM-0.2M NaCl. These results suggest that the influence of salt concentration on the velocity of aminoacylation depends on both the conformational changes of tRNA and seryl-tRNA synthetase.     

Changes in serum testosterone and estradiol concentrations following acute androstenedione ingestion in young women.

The effect of androstenedione intake on serum hormone concentrations in women is equivocal. Therefore, we examined the hormonal response to androstenedione intake in healthy young (22.1 +/- 0.4 y) women for 4 hours. On day 3 of the follicular phase, subjects ingested placebo, 100, or 300 mg androstenedione in a random, double-blind, cross-over manner. Blood samples were collected before and every 30 min for 240 min after intake. Serum androstenedione concentrations (means +/- SE) increased above basal (6.2 +/- 0.8 nmol/l) from 60-240 min for both 100 mg (22.6 +/- 1.0 nmol/l at 240 min) and 300 mg (28.1 +/- 1.3 nmol/l at 210 min). Androstenedione intake increased serum total testosterone concentrations above basal (1.2 +/- 0.2 nmol/l) from 120-240 min (5.5 +/- 0.9 nmol/l at 210 min) with 100 mg and from 60-240 with 300 mg (10.2 +/- 1.6 nmol/l at 210 min). Androstenedione intake also increased serum estradiol concentrations (basal 191 +/- 24 pmol/l) at 150 min with 100 mg (237 +/- 35 pmol/l) and from 150-240 min with 300 mg (reaching 260 +/- 32 pmol/l at 240 min). These data indicate that, in contrast to men, androstenedione intake in women increases serum testosterone concentrations.     

Effect of anesthetics on the stability of oxyhemoglobin S

The conversion of the serine-195 in α-chymotrypsin to dehydroalanine results in two conformational substates that differ in their extinction coefficients at 240nm. The active site methionine-192 in the substate with lower absorption at 240nm is alkylated by α-bromo-4-nitroacetophenone at a rate of 7.0×10^?4sec^?1, similar to that found for α-chymotrypsin; the substate with higher absorption at 240nm reacts 14 times slower. These two substates are not separated by an affinity resin containing lima bean trypsin inhibitor. These data infer that the serine-195 plays a role in the stabilization of the active site conformation in α-chymotrypsin.     

Temperature dependence of the optical activity of human serum low density lipoprotein. The role of lipids.

Low density lipoprotein (LDL) (1.024-1.045 G/cm3) was prepared by ultracentrifugal flotation from serum of normal fasting subjects. Circular dichroism (CD) and optical rotatory dispersion (ORD) spectra in the ultraviolet region were measured at 2, 25, and 37 degrees on LDL, lipid extracted from LDL, and on pure component lipids. All exhibit reversible, temperature-dependent optical activities. Sphingomyelin has a strong negative CD band around 195 nm. Cholesterol and cholesteryl esters have a CD minimum at 208 nm. They have positive CD bands around 201 and 198 nm which decrease sharply and become negative at 198 and 193 nm, respectively. The CD of the total lipid extract of LDL is negative and drops monotonically below 200 nm. Thus, the lipid moiety could account for the increasing negativity of the CD of LDL below 195 nm. After subtraction of the ellipticity corresponding to amounts of lipids in organic solvents equivalent to those found in LDL, the 208-210 nm trough of LDL diminishes markedly. This is accompanied by a blue-shift of the extrema from 195-196 to 193 nm and an increase in the magnitude of the positive ellipticity. The fractions of helix and of beta form in the protein, determined by the method of Y. H. Chen, J. T. Yang, and K. H. Chau ((1974), Biochemistry 13, 3350), in the wavelength interval of 250-240 nm, remain essentially unchanged between 2 and 37 degrees. These observations suggest that a substantial part of the thermal change in the CD spectrum of LDL between 208 and 210 nm may be attributable to lipids.     

The time dependent UV resonance Raman spectra, conformation, and biological activity of acetylcholine analogues upon binding to acetylcholine binding proteins.

In order to obtain quantitative data on the relation between the conformation of acetylcholine and its interaction with biologically significant proteins, a series of acetylcholine analogues with absorption bands in the region 200-300 nm have been synthesized or obtained commercially. Each of these compounds were assayed to measure its activity as an ion channel activator of the nicotinic acetylcholine receptor protein (AChR). In addition, the suitability of some of these compounds as substrates for hydrolysis by acetylcholine esterase (AChE) was determined. One of these analogues, dimethylthionocarbamylcholine (DMTC-Ch), has the ester carbonyl oxygen replaced by a thionyl sulfur. DMTC-Ch has been found to be quite active as an ion channel activator when bound to AChR and was found to react with the enzyme AChE as a suicide substrate. It forms a thionoester of the serine at the AChE active site by an ester exchange reaction that releases the choline as the first product. However, the second or acid product is not released even at pH 7.5 over a period of days. This acetylcholine analog has an absorption band at about 240 nm and exhibits very strong ultraviolet resonance Raman (UVRR) spectra using 239 nm excitation from a frequency modified Nd:YAG laser. This technique allows observation of both conformational changes of the ligand molecule that result in frequency changes as well as changes in the excited state electronic structure that results in changes in the relative intensity of the Raman bands. The time dependence of the UVRR spectrum of the ligand upon binding to both AChE and AChR has been studied from 0.1 msec to minutes. Some time dependence in the conformation of DMTC-Ch upon binding to AChE has been found for very short (0.1-0.5 msec) times. However, no change in the conformation of this neurotransmitter analog is found in the available time range upon binding to AChR. From these data it is concluded that a previous suggestion that acetylcholine has a conformational change upon binding to AChR may be incorrect since the solution behavior of the carbamyl cholines and acetylcholine are similar. Even if acetylcholine does change conformation upont binding to AChR, it is unlikely that such a conformational change plays a significant role in channel activation. We present strong evidence that acetylcholine and its analogues can be active in a variety of conformations.(ABSTRACT TRUNCATED AT 400 WORDS)     

Far-ultraviolet optical activity of saccharide derivatives. II. Dextran,amylose, and mycodextran acetes; dextran and amylose xanthates

The ultraviolet ORD and CD spectra of amylose, dextran, and mycodextran acetates and some of thier oligomers were recorded in trifluoroethanol solution in the 300–185nm wavelength range. Similarly, the spectra of amylose and dextran xanthates in water solution were obtained in the 400–200 nm range. In the amylose acetate series, the monomer and dimer both show a normal acetyl n → π* transition in CD, while the trimer and the polymer both exhibit an additional, shorter wavelength peak. The latter is presumed to arise from a helical conformation of the amylose chain. This interpretation is substantiated by a helix–coil type transition of the CD spectra of amylose triacetate at elevated temperatures and a reversion of the anomalous CD to the normal CD upon partial deacetylation. By contrast, neither dextran acetates nor mycodextran acetate exhibit any conformational effects. The CD of dextran acetates is quite sensitive to β-1,6 and branch linkages. The ORD and CD of amylose xanthate are complex, suggesting the presence of organized structure in solution. The dextran xanthate shows only a simple ORD spectrum and no observable CD.     

Small angle x-ray study on the structure of active and inactive ribulose bisphosphate carboxylase from Alcaligenes eutrophus. Evidence for a configurational change

Two small angle x-ray scattering curves have been obtained from active and inactive ribulose 1,5-bisphosphate carboxylase from Alcaligenes eutrophus. The radius of gyration was calculated to be R = 47.8 +/- 0.1 nm for the active enzyme and R = 49.2 +/- 0.1 nm for the inactive enzyme. The maximum particle dimension amounts to 13.5 +/- 0.5 nm for the active and 15.7 +/- 0.5 nm for the inactive enzyme. A model of the active carboxylase is presented. It is in good agreement with models derived from electron microscopical data. Model calculations for the inactive enzyme show some evidence for a configurational change.     

Molecular Dynamic Simulations of the N-Terminal Receiver Domain of NtrC Reveal Intrinsic Conformational Flexibility in the Inactive State

Abstract

The N-terminal receiver domain of NtrC is the molecular switch in the two-component signal transduction. It is the first protein where structures of both the active (phosphyroylated) and inactive (unphosphyroylated) states are determined experimentally. Phosphorylation of the NtrC at the active site induces large structural change. NMR experiments suggested that the wild type unphosphorylated NtrC adopts both the active and the inactive conformations and the phosphorylation stabilizes the active conformations. We applied free (unconstrained) molecular dynamic (MD) simulation to examine the intrinsic flexibilities and stabilities of the NtrC receiver domain in both the active and inactive conformations. Molecular dynamic simulations showed that the inactive state of NtrC receiver domain is more flexible than the active state. There were large movements in helix 4 and loop β3-α3 which coincide with major structural differences between the inactive and active states. We observed large root-mean-square deviations from the initial starting structure and the large root-mean-square fluctuations during MD simulation for the inactive state. We then investigated the activation pathway with Targeted MD simulation. We show that the intrinsic flexibility in the loop β3-α3 plays an important role in triggering the conformational change. Phosphorylation at the active site may serve to stabilize the conformational change. These results together suggest that the unphosphorylated NtrC receiver domain could be involved in a conformational equilibrium between two different states.     

Allosteric modulator ORG27569 induces CB1 cannabinoid receptor high affinity agonist binding state, receptor internalization, and Gi protein-independent ERK1/2 kinase activation

The cannabinoid receptor 1 (CB1), a member of the class A G protein-coupled receptor family, is expressed in brain tissue where agonist stimulation primarily activates the pertussis toxin-sensitive inhibitory G protein (G(i)). Ligands such as CP55940 ((1R,3R,4R)-3-[2-hydroxy-4-(1,1-dimethylheptyl)phenyl]-4-(3- hydroxypropyl)cyclohexan-1-ol) and Δ(9)-tetrahydrocannabinol are orthosteric agonists for the receptor, bind the conventional binding pocket, and trigger G(i)-mediated effects including inhibition of adenylate cyclase. ORG27569 (5-chloro-3-ethyl-1H-indole-2-carboxylic acid [2-(4-piperidin-1-yl-phenyl)ethyl]amide) has been identified as an allosteric modulator that displays positive cooperativity for CP55940 binding to CB1 yet acts as an antagonist of G protein coupling. To examine this apparent conundrum, we used the wild-type CB1 and two mutants, T210A and T210I (D'Antona, A. M., Ahn, K. H., and Kendall, D. A. (2006) Biochemistry 45, 5606-5617), which collectively cover a spectrum of receptor states from inactive to partially active to more fully constitutively active. Using these receptors, we demonstrated that ORG27569 induces a CB1 receptor state that is characterized by enhanced agonist affinity and decreased inverse agonist affinity consistent with an active conformation. Also consistent with this conformation, the impact of ORG27569 binding was most dramatic on the inactive T210A receptor and less pronounced on the already active T210I receptor. Although ORG27569 antagonized CP55940-induced guanosine 5'-3-O-(thio)triphosphate binding, which is indicative of G protein coupling inhibition in a concentration-dependent manner, the ORG27569-induced conformational change of the CB1 receptor led to cellular internalization and downstream activation of ERK signaling, providing the first case of allosteric ligand-biased signaling via CB1. ORG27569-induced ERK phosphorylation persisted even after pertussis toxin treatment to abrogate G(i) and occurs in HEK293 and neuronal cells.     

Near- and far-ultraviolet circular dichroism of the catalytic subunit of adenosine cyclic 5'-monophosphate dependent protein kinase

The circular dichroism spectrum of the catalytic subunit of cAMP-dependent protein kinase was measured in the far-UV (190-240 nm) and near-UV (250-300 nm) region. Data from the far-UV spectra were processed with the CONTIN program for estimation of globular protein secondary structure [ Provencher , S. W. (1982) CONTIN (Version 2) User's Manual, European Molecular Biology Laboratory, Heidelberg, West Germany]. The composition of the protein determined by this method was 49 +/- 2% alpha-helix, 20 +/- 4% beta-sheet, and 31 +/- 3% remainder. This composition changes when the protein is allowed to bind Kemptide , a synthetic peptide substrate, with more than half of the disordered portion of the protein taking the form of beta-sheet. A certain portion of the alpha-helical structure also appears to move into a beta-sheet form. The near-UV CD spectrum of catalytic subunit shows changes in aromatic amino acid dichroism associated with substrate binding. These changes can be ascribed with a fair degree of certainty to alterations in the orientation of a tyrosine residue at the surface of the protein. These findings are discussed in terms of previous work on induced dichroism in this enzyme with regard to control mechanisms operating at the active site.     

D1-D2-cytochrome b559 complex from the aquatic plant Spirodela oligorrhiza: correlation between complex integrity, spectroscopic properties, photochemical activity, and pigment composition

A D1-D2-cyt b559 complex with about four attached chlorophylls and two pheophytins has been isolated from photosystem II of the aquatic plant Spirodela oligorrhiza and used for studying the detergent-induced changes in spectroscopic properties and photochemical activity. Spectral analyses (absorption, CD, and fluorescence) of D1-D2-cyt b559 preparations that were incubated with different concentrations of the detergent Triton X-100 indicate two forms of the D1-D2-cyt b559 complexes. One of them is photochemically active and has an absorption maximum at 676 nm, weak fluorescence at 685 nm, and a strong CD signal. The other is photochemically inactive, with an absorption maximum at 670 nm, strong fluorescence at 679 nm, and much weaker CD. The relative concentrations of the two forms determine the overall spectra of the D1-D2-cyt b559 preparation and can be deduced from the wavelength of the lowest energy absorption band: preparations having maximum absorption at 674, 672, or 670.5 nm have approximately 20, 60, or 85% inactive complexes. The active form contains two chlorophylls with maximum absorption at 679 nm and CD signals at 679 (+) and 669 nm (-). These chlorophylls make a special pair that is identified as the primary electron donor P-680. The calculated separation between the centers of these two pigments (using an extended version of the exciton theory) is about 10 A, the pigments' molecular planes are tilted by about 20 degrees, and their N1-N3 axes are rotated by 150 degrees relative to each other. The other two chlorophylls and one of the two pheophytins in the D1-D2-cyt b559 complex have their maximum absorption at 672 nm, while the maximum absorption of the photochemically active pheophytin is probably at 672-676 nm. During incubation with Triton X-100, the photochemically active complex is transformed into an inactive form with first-order kinetics. In the inactive form the maximum absorption of the 679 nm absorbing Chls is blue-shifted to 669 nm. The first-order decay of the photochemical activity suggests that the isolated D1-D2-cyt b559 complex is stable as an aggregate but becomes unstable on dissociation into individual D1-D2-cyt b559 units.     

Physico-chemical properties of deoxyribonucleoprotamine from sturgeon sperm heads]

Deoxyribonucleoprotamine (DNPn) from sonicated nuclei of sturgeon sperm heads was studied by means of ring dichroism. A derivative analysis of DNA and DNPn melting curves in 1 mM Tris. HCl pH 8.0 revealed the fraction of protein-free DNA being about 30% and suggested the preferable binding of protamine molecules with AT-rich DNA regions. The latter is also confirmed by the data on ring dichroism of protein-poor soluble DNAPn fraction in 0,14 M NaCl. Ring dichroism of DNA and DNPn in 1 mM Tris coinsides at the wavelength of 310-240 nm at concentrations of 500-50 mkg/ml. Dilution of DNPn to 5 mkg-ml resulted in the decrease of the ellipticity at 275 nm and produced no effect at 260-210 nm. The effect observed is suggested to be due to a partial transition of DNA in DNPn into C-form under the dilution as a result of a higher molecule hydration and a destruction of some hydrogen bonds between guanidine residues of arginine and oxygen of phosphate groups, stabilyzing DNA in the B-form. Ring dichroism spectrum of protamine, calculated by the subtraction of DNA spectrum from DNPn spectrum at the region of 240-210 nm coinsides with that of free protamine and indicates the absence of an ordered structure in protamine molecules in DNPn.     

Molecular dynamic simulations of the N-terminal receiver domain of NtrC reveal intrinsic conformational flexibility in the inactive state

The N-terminal receiver domain of NtrC is the molecular switch in the two-component signal transduction. It is the first protein where structures of both the active (phosphyroylated) and inactive (unphosphyroylated) states are determined experimentally. Phosphorylation of the NtrC at the active site induces large structural change. NMR experiments suggested that the wild type unphosphorylated NtrC adopts both the active and the inactive conformations and the phosphorylation stabilizes the active conformations. We applied free (unconstrained) molecular dynamic (MD) simulation to examine the intrinsic flexibilities and stabilities of the NtrC receiver domain in both the active and inactive conformations. Molecular dynamic simulations showed that the inactive state of NtrC receiver domain is more flexible than the active state. There were large movements in helix 4 and loop beta3-alpha3 which coincide with major structural differences between the inactive and active states. We observed large root-mean-square deviations from the initial starting structure and the large root-mean-square fluctuations during MD simulation for the inactive state. We then investigated the activation pathway with Targeted MD simulation. We show that the intrinsic flexibility in the loop beta3-alpha3 plays an important role in triggering the conformational change. Phosphorylation at the active site may serve to stabilize the conformational change. These results together suggest that the unphosphorylated NtrC receiver domain could be involved in a conformational equilibrium between two different states.     

Agonist-induced conformational changes in bovine rhodopsin: insight into activation of G-protein-coupled receptors

Activation of G-protein-coupled receptors (GPCRs) is initiated by conformational changes in the transmembrane (TM) helices and the intra- and extracellular loops induced by ligand binding. Understanding the conformational changes in GPCRs leading to activation is imperative in deciphering the role of these receptors in the pathology of diseases. Since the crystal structures of activated GPCRs are not yet available, computational methods and biophysical techniques have been used to predict the structures of GPCR active states. We have recently applied the computational method LITiCon to understand the ligand-induced conformational changes in β₂-adrenergic receptor by ligands of varied efficacies. Here we report a study of the conformational changes associated with the activation of bovine rhodopsin for which the crystal structure of the inactive state is known. Starting from the inactive (dark) state, we have predicted the TM conformational changes that are induced by the isomerization of 11-cis retinal to all-trans retinal leading to the fully activated state, metarhodopsin II. The predicted active state of rhodopsin satisfies all of the 30 known experimental distance constraints. The predicted model also correlates well with the experimentally observed conformational switches in rhodopsin and other class A GPCRs, namely, the breaking of the ionic lock between R135^3.50 at the intracellular end of TM3 (part of the DRY motif) and E247^6.30 on TM6, and the rotamer toggle switch on W265^6.48 on TM6. We observe that the toggling of the W265^6.48 rotamer modulates the bend angle of TM6 around the conserved proline. The rotamer toggling is facilitated by the formation of a water wire connecting S298^7.45, W265^6.48 and H211^5.46. As a result, the intracellular ends of TMs 5 and 6 move outward from the protein core, causing large conformational changes at the cytoplasmic interface. The predicted outward movements of TM5 and TM6 are in agreement with the recently published crystal structure of opsin, which is proposed to be close to the active-state structure. In the predicted active state, several residues in the intracellular loops, such as R69, V139^3.54, T229, Q237, Q239, S240, T243 and V250^6.33, become more water exposed compared to the inactive state. These residues may be involved in mediating the conformational signal from the receptor to the G protein. From mutagenesis studies, some of these residues, such as V139^3.54, T229 and V250^6.33, are already implicated in G-protein activation. The predicted active state also leads to the formation of new stabilizing interhelical hydrogen-bond contacts, such as those between W265^6.48 and H211^5.46 and E122^3.37 and C167^4.56. These hydrogen-bond contacts serve as potential conformational switches offering new opportunities for future experimental investigations. The calculated retinal binding energy surface shows that binding of an agonist makes the receptor dynamic and flexible and accessible to many conformations, while binding of an inverse agonist traps the receptor in the inactive state and makes the other conformations inaccessible.     

Randomization of the entire active-site helix alpha 1 of the thiol-disulfide oxidoreductase DsbA from Escherichia coli

DsbA from Escherichia coli is the most oxidizing member of the thiol-disulfide oxidoreductase family (E(o)' = -122 mV) and is required for efficient disulfide bond formation in the periplasm. The reactivity of the catalytic disulfide bond (Cys(30)-Pro(31)-His(32)-Cys(33)) is primarily due to an extremely low pK(a) value (3.4) of Cys(30), which is stabilized by the partial positive dipole charge of the active-site helix alpha1 (residues 30-37). We have randomized all non-cysteine residues of helix alpha1 (residues 31, 32, and 34-37) and found that two-thirds of the resulting variants complement DsbA deficiency in a dsbA deletion strain. Sequencing of 98 variants revealed a large number of non-conservative replacements in active variants, even at well conserved positions. This indicates that tertiary structure context strongly determines alpha-helical secondary structure formation of the randomized sequence. A subset of active and inactive variants was further characterized. All these variants were more reducing than wild type DsbA, but the redox potentials of active variants did not drop below -210 mV. All inactive variants had redox potentials lower than -210 mV, although some of the inactive proteins were still re-oxidized by DsbB. This demonstrates that efficient oxidation of substrate polypeptides is the crucial property of DsbA in vivo.     

Structural conversion between open and closed forms of radixin: low-angle shadowing electron microscopy

The function of ERM (ezrin/radixin/moesin) proteins as general cross-linkers between actin filaments and plasma membranes is regulated downstream of Rho, through the transition between active and inactive forms. To directly examine the conformational change between the active and inactive forms of ERM proteins, we applied low-angle rotary-shadowing electron microscopy to the radixin molecules, wild-type, T564A-non-phosphorylated-type, and T564E-phosphorylated-type, since most of the active forms are reportedly stabilized in cells by the C-terminal threonine phosphorylation. As a result, the T564A- and wild-type radixin molecules yielded the globular closed forms, approximately 8-14 nm in diameter, with some striations on their surfaces. In contrast, the T564E-radixin molecules tended to take elongated open forms, in which two globular structures measuring approximately 8 nm and approximately 5 nm in diameter were associated with both ends of the filamentous structures. The filamentous structure took either a approximately 20-25 nm-long straight course or a folded course. Taken together with the biochemical and the crystal structural results obtained to date, the closed and open forms represent the inactive and active forms of radixin as cross-linkers between actin filaments and plasma membranes.     

Study of the structure of the histidine decarboxylase of Micrococcus sp. n. by the method of circular dichroism]

Ring dichroism spectra (RD) of histidine decarboxylase (HDC) from Micrococcus sp. n. at the regions of peptide bonds (200-240 nm) and aromatic amino acids (250-300 nm) absorption are studied. The treatment of RD spectra according to methods of Greenfield-Fasman, Saksena-Vetlaufer and Mayer permits to conclude that at the pH range within 4-8 the content of ordered structures of alpha-helix type comprises 20%, that of beta-structure type-40%, while the rest 40% are represented with polypeptide chain in a disordered globular state. When pH is varied from 1 to 12, the content of alpha-helices decreases from 17 to 5%. There are two distinct dichroic bands in the spectrum of aromatic chromophores absorption (at 270 and 290 nm), the former containing tirosine, tryptophane and phenylalanine residues and the latter being induced with triptophane residues. The study of HDC RD spectra at the regions of peptide bonds and aromatic acids absorption at different temperatures has shown that a part of triptophane, tyrosine and phenylalanine residues is in an ordered structure of the alpha-helix type. The HDC undergoes irreversible changes under heating to 70 degrees and in 8 M urea. 5 M guanidine chloride eliminates the ordered HDC structure, while sodium dodecylsulphate at concentrations up to 1% does not affect the enzyme structure.     

2013年北太平洋公海渔场柔鱼体内典型放射性核素分析及风险评估

为了解福岛核泄漏放射性核素对北太平洋公海渔场的后续性污染影响情况,于2013年8—11月在北太平洋公海渔场(40°—44° N、145°—160° E)采集柔鱼样品,利用超低本底HPGe γ 谱仪分析5种放射性核素.结果表明: ¹³⁴Cs和^110mAg未检测出高于本底值的比活度,而²¹⁰Pb、¹³⁷Cs和⁴⁰K均有检出.其中²¹⁰Pb在柔鱼体的比活度范围为0.99～7.60 Bq·kg^-1,¹³⁷Cs的比活度范围为最低检测限～0.37 Bq·kg^-1,⁴⁰K的比活度范围为46.00～107.00 Bq·kg^-1.²¹⁰Pb核素含量在柔鱼250～300 mm胴长组分布最高;在200～250和>400 mm胴长组中最低.¹³⁷Cs含量在柔鱼200～250 mm胴长组中最高;在250～300和350～400 mm胴长组较低.⁴⁰K核素含量在柔鱼250～300 mm胴长组中最高,在150～200 mm胴长组最低,总体分布比较均匀.3种核素在柔鱼的不同组织中含量不同,但均在内脏中含量最高.通过风险评估发现,北太平洋公海渔场所有检测的海洋生物的放射性核素含量值与基准值的熵值均未超过1,说明目前核辐射对该海域渔业资源的影响暂无大风险.研究可为福岛核事故对海洋环境生态影响及公众健康危害评估提供理论依据.     

Mutations of CB1 T210 produce active and inactive receptor forms: correlations with ligand affinity, receptor stability, and cellular localization

Human cannabinoid receptor 1 (CB(1)) has attracted substantial interest as a potential therapeutic target for treating obesity and other obsessive disorders. An understanding of the mechanism governing the transition of the CB(1) receptor between its inactive and active states is critical for understanding how therapeutics can selectively regulate receptor activity. We have examined the importance of the Thr at position 210 in CB(1) in this transition, a residue predicted to be on the same face of the helix as the Arg of the DRY motif highly conserved in the G protein-coupled receptor superfamily. This Thr was substituted with Ile and Ala via mutagenesis, and the receptors, T210I and T210A, were expressed in HEK 293 cells. The T210I receptor exhibited enhanced agonist and diminished inverse agonist affinity relative to the wild type, consistent with a shift toward the active form. However, treatment with GTPgammaS to inhibit G protein coupling diminished the affinity change for the inverse agonist SR141716A. The decreased thermal stability of the T210I receptor and increased level of internalization of a T210I receptor-GFP chimera were also observed, consistent with constitutive activity. In contrast, the T210A receptor exhibited the opposite profile: diminished agonist and enhanced inverse agonist affinity. The T210A receptor was found to be more thermally stable than the wild type, and high levels of a T210A receptor-GFP chimera were localized to the cell surface as predicted for an inactive receptor form. These results suggest that T210 plays a key role in governing the transition between inactive and active CB(1) receptor states.