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1.
The large-conductance, voltage- and Ca2+-gated K+ (BK) channel consists of four α subunits, which form a voltage- and Ca2+-gated channel, and up to four modulatory β subunits. The β1 subunit is expressed in smooth muscle, where it slows BK channel kinetics and shifts the conductance–voltage (G-V) curve to the left at [Ca2+] > 2 µM. In addition to the six transmembrane (TM) helices, S1–S6, conserved in all voltage-dependent K+ channels, BK α has a unique seventh TM helix, S0, which may contribute to the unusual rightward shift in the G-V curve of BK α in the absence of β1 and to a leftward shift in its presence. Such a role is supported by the close proximity of S0 to S3 and S4 in the voltage-sensing domain. Furthermore, on the extracellular side of the membrane, one of the two TM helices of β1, TM2, is adjacent to S0. We have now analyzed induced disulfide bond formation between substituted Cys residues on the cytoplasmic side of the membrane. There, in contrast, S0 is closest to the S2–S3 loop, from which position it is displaced on the addition of β1. The cytoplasmic ends of β1 TM1 and TM2 are adjacent and are located between the S2–S3 loop of one α subunit and S1 of a neighboring α subunit and are not adjacent to S0; i.e., S0 and TM2 have different trajectories through the membrane. In the absence of β1, 70% of disulfide bonding of W43C (S0) and L175C (S2–S3) has no effect on V50 for activation, implying that the cytoplasmic end of S0 and the S2–S3 loop move in concert, if at all, during activation. Otherwise, linking them together in one state would obstruct the transition to the other state, which would certainly change V50.  相似文献   

2.
Summary The secondary structure of 5S rRNA has been elucidated by a cladistic analysis resulting in minimal models for eukaryotes, eubacteria, and halophilic-methanogenic archaebacteria, as well as for an ur-5S rRNA. This ancestor of all present-day 5S rRNA molecules is compared with an ur-tRNA and can be fitted into a tRNA-like structure allowing tertiary-structure interactions at the equivalent positions. A phylogenetic analysis of eukaryotic 5SrRNA and 16S rRNA sequences confirms particular monophyletic taxa: rhodophytes (red algae), chlorobionts (green algae and plants), metazoans (multicellular animals), euglenozoans (euglenids and trypanosomatids), a group of zygomycetes (excluding Kickxellales), a group of ascomycetes (excluding Protomycetales), two distinct groups of basidiomycetes, and a group consisting of phaeophyceans (brown algae) and oomycetes (water molds). The Euglenozoa show a distinct relation to the Eumycota (true fungi) and Metazoa. An analysis of archaebacterial sequences substantiates the paraphyletic nature of this third urkingdom defining the eubacteria as a sister group of the halophile-methanogens and defining the eukaryotes as a sister group of a particular lineage of the eocytes/sulfur-dependents. The latter fact implies that even the eocytes/sulfur-dependent archaebacteria are paraphyletic.Presented at the FEBS Symposium on Genome Organization and Evolution, held in Crete, Greece, September 1–5, 1986Dedicated to the memory of Erik Huysmans who died on July 8, 1986, at the age of 29.  相似文献   

3.
Rubredoxins contain a mononuclear iron tetrahedrally coordinated by four cysteinyl sulfurs. We have studied the wild-type protein from Clostridium pasteurianum and two mutated forms, C9S and C42S, in the oxidized and reduced states, with Mössbauer, integer-spin EPR, and magnetic circular dichroism (MCD) spectroscopies. The Mössbauer spectra of the ferric C42S and C9S mutant forms yielded zero-field splittings, D=1.2?cm?1, that are about 40% smaller than the D-value of the wild-type protein. The 57Fe hyperfine coupling constants were found to be ca. 8% larger than those of the wild-type proteins. The present study also revealed that the ferric wild-type protein has δ=0.24±0.01?mm/s at 4.2?K rather than δ=0.32?mm/s as reported in the literature. The Mössbauer spectra of both dithionite-reduced mutant proteins revealed the presence of two ferrous forms, A and B. These forms have isomer shifts δ=0.79?mm/s at 4.2?K, consistent with tetrahedral Fe2+(Cys)3(O-R) coordination. The zero-field splittings of the two forms differ substantially; we found D=?7±1?cm?1, E/D=0.09 for form A and D=+6.2±1.3?cm?1, E/D=0.15 for form B. Form A exhibits a well-defined integer-spin EPR signal; from studies at X- and Q-band we obtained g z =2.08±0.01, which is the first measured g-value for any ferrous rubredoxin. It is known from X-ray crystallographic studies that ferric C42S rubredoxin is coordinated by a serine oxygen. We achieved 75% reduction of C42S rubredoxin by irradiating an oxidized sample at 77?K with synchrotron X-rays; the radiolytic reduction produced exclusively form A, suggesting that this form represents a serine-bound Fe2+ site. Studies in different buffers in the pH?6–9 range showed that the A:B ratios, but not the spectral parameters of A and B, are buffer dependent, but no systematic variation of the ratio of the two forms with pH was observed. The presence of glycerol (30–50% v/v) was found to favor the B form. Previous absorption and circular dichroism studies of reduced wild-type rubredoxin have suggested d-d bands at 7400, 6000, and 3700?cm?1. Our low-temperature MCD measurements place the two high-energy transitions at ca. 5900 and 6300?cm?1; a third d-d transition, if present, must occur with energy lower than 3300?cm?1. The mutant proteins have d-d transitions at slightly lower energy, namely 5730, 6100?cm?1 in form A and 5350, 6380?cm?1 in form B.  相似文献   

4.
At present the majority of the endemic bird species occurring on São Tomé and Príncipe remain common, the rarer species being those largely confined to primary rainforest. In 1990 the dwarf olive ibis (Bostrychia bocagei), São Tomé fiscal shrike (Lanius newtoni), São Tomé short-tail (Amaurocichla bocagii) were rediscovered and in 1991 the São Tomé grosbeak (Neospiza concolor) was seen for the first time since 1888. Lowland primary forest is the only habitat on São Tomé in which all the endemic species are found. Primary forest on Príncipe remains largely unsurveyed since the beginning of the century. Due to the decline in the cocoa industry and poor infra-structure post-independence the extent of secondary forest is probably at its greatest since the late 1800s. This habitat is an important buffer zone against development for the remaining primary forest and also contains important populations of many of the endemic species, in particular the São Tomé scops owl (Otus hartlaubi), São Tomé oriole (Oriolus crassirostris) and giant weaver (Ploceus grandis). On both islands the remaining areas of primary forest need immediate protection and suitable boundaries have been designated under the Zona Ecológica plan. Fortunately, except for an important area around Lagôa Amélia, primary forest is not under immediate threat, although a variety of pressures are likely to increase. In conjunction with the protection of primary forest, a plant for managing the remaining timber resource on the islands will be a vital requirement, particularly if areas of shade forest, an important habitat for endemic bird species and a potentially valuable economic resource, are to be conserved and sustainably managed.  相似文献   

5.

Aim

To evaluate the current treatment of mycosis fungoides (MF) and Sézary syndrome (SS) focusing on the role of radiotherapy (RT), its principles and indications, and the perspectives of the novel irradiation technologies.

Background

MF and SS are rare lymphoproliferative diseases whose incidence is increasing. For a long time RT has been used as a single modality or in integrated treatment programs for these diseases.

Materials and methods

The latest systematic reviews, primary studies and new diagnostic and treatment guidelines on MF and SS were analyzed. Clinical outcomes together with the technical aspects and the role of RT were also evaluated.

Results

New data are available on pathogenesis, diagnostic criteria, classification and staging procedures for MF and SS and several local and systemic therapies are proposed. Localized RT can cure “minimal stage” MF while total skin electron beam irradiation (TSEI) may cure initial-stage disease and may offer important symptom relief (itch, erythroderma) in a more advanced setting. Despite its efficacy, RT is not largely used, mainly because of some technical difficulties but new RT technologies may be proposed to treat large skin surfaces.

Conclusions

New treatment programs offer good results, with median survival of more than 12 years in early-stage MF, but the median survival of 2.5 years or less in advanced stages is still a challenge. RT remains an option for all stages with a good cost/effectiveness ratio in a curative or palliative setting. New RT technologies can overcome some technical problems of treating large skin surfaces.  相似文献   

6.
Quinoxaline-2-carboxylic acid (QXC) and 3-hydroxyquinaldic acid (HQA) feature in quinomycin family and confer anticancer activity. In light of the significant potency against cancer, the biosynthetic gene clusters have been reported from many different Streptomyces strains, and the biosynthetic pathway were proposed mainly based on the in vivo feeding experiment with isotope labeled putative intermediates. Herein we report another gene cluster from Streptomyces griseovariabilis subsp. bandungensis subsp. nov responsible for the biosynthesis of echinomycin (a member of quinomycin family, also named quinomycin A) and presented in vitro evidence to corroborate the previous hypothesis on QXC biosynthesis, showing that only with the assistance of a MbtH-like protein Qui5, did the didomain NRPS protein (Qui18) perform the loading of a L-tryptophan onto its own PCP domain. Particularly, it was found that Qui5 and Qui18 subunits form a functional tetramer through size exclusion chromatography. The subsequent hydroxylation on β-carbon of the loaded L-tryptophan proved in vitro to be completed by cytochrome P450-dependent hydroxylase Qui15. Importantly, only the Qui18 loaded L-tryptophan can be hydroxylated by Qui15 and the enzyme was inactive on free L-tryptophan. Additionally, the chemically synthesized (2S,3S) β-hydroxytryptophan was detected to be converted by the tryptophan 2,3-dioxygenase Qui17 through LC-MS, which enriched our previous knowledge that tryptophan 2,3-dioxygenase nearly exclusively acted on L-tryptophan and 6-fluoro-tryptophan.  相似文献   

7.
Summary The stereoselectivity of penicillin amidase (PA, EC 3.5.1.11) from E coli and homologeous enzymes from other sources has been determined as a function of temperature and substrate for hydrolysis and kinetically controlled synthesis. The stereoselectivity of these reactions decreased almost by one order of magnitude from 5 to 45°C. It increased with the substrate (k cat/K m) and nucleophile (k T/k H) specificity, and was found to differ in the S1- (R-specific) and S1-(S-specific)-binding subsites of the active site. The S1-stereoselectivity was determined mainly by differences in the activation energy, i.e. the turnover number. The stereoselectivity of PA from different sources differed by almost an order of magnitude for the same substrate.  相似文献   

8.
Streptomyces lividans IAF18, obtained by homologous cloning, is capable of over-producing XlnA. To investigate the possibility of the expression of foreign genes, various coding regions of the xylanase A gene (xlnA) were analysed. Expression/secretion vectors were constructed containing the regulatory elements of xlnA with the coding region of the leader peptide with or without the truncated structural gene encoding the first 310 amino acids of the XlnA. The genes coding for the Escherichia coliβ-glucuronidase and subunit 1 of the Bordetella pertussis toxin (S1) were used and their expression analysed. S. lividans transformants where the β-glucuronidase gene was fused with the leader sequence produced up to 30 mg β-glucuronidase/culture filtrate whereas only fused XlnA/S1 was detected and its yield was estimated to be 1 mg/l. The disappearence of the B. pertussis toxin S1 and β-glucuronidase from the culture medium was due to the concomitant appearence of secreted proteases from S. lividans. Received: 19 July 1995/Received revision: 3 November 1995/Accepted: 20 November 1995  相似文献   

9.
3′–nucleases/nucleotidases of the S1–P1 family (EC 3.1.30.1) are single–strand–specific or non-specific zinc–dependent phosphoesterases present in plants, fungi, protozoan parasites, and in some bacteria. They participate in a wide variety of biological processes and their current biotechnological applications rely on their single–strand preference, nucleotide non-specificity, a broad range of catalytic conditions and high stability. We summarize the present and potential utilization of these enzymes in biotechnology and medicine in the context of their biochemical and structure–function properties. Explanation of unanswered questions for bacterial and trypanosomatid representatives could facilitate development of emerging applications in medicine.  相似文献   

10.
Summary A new strategy for stereoconservative and stereoselective syntheses of several types of amino acids starting from-functional carboxylic acids employing hexafluoroacetone as protecting and activating reagent is described. Outstanding features of this new method are the mild reaction conditions and the high yields for introduction and cleavage of the protective group allowing sensitive functional groups in the side chain to survive. Furthermore, the new concept results in saving of synthetic steps.  相似文献   

11.
Summary Plasma membranes were prepared from soybean hypocotyls and roots by aqueous two-phase partitioning and subsequent free-flow electrophoresis. The highly purified plasma membranes bound [35S]GTPS with a relatively high affinity (Kd10nM). The binding was saturable and specific as it was indicated by the displacement of bound [35S]GTPS by unlabeled GTPS and GTP, but not by ATPS, ATP, UTP or CTP. ITP was intermediate in its ability to displace [35S]GTPS. When soybean plasma membrane proteins were separated by SDS-PAGE and displayed by autoradiography, two major [35S]GTPS binding proteins were revealed with apparent molecular weights of 24 and 28 kDa. Results with plasma membranes from soybean hypocotyls and roots were similar but differed from those with plasma membranes prepared from rat liver and adipocytes where only a single major [35S]GTPS binding activity with a molecular weight of 28 kDa was observed.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - G protein hetero-trimeric GTP binding protein with , , subunits - Gn protein GTP binding protein detected on nitrocellulose blots - GTPS guanosine 5-[-thio]triphosphate - IAA 3-indoleacetic acid - SDS-PAGE sodium dodecylsulfate-polyacrylamide gel electrophoresis  相似文献   

12.
The pore forming toxin Hla (α-toxin) from Staphylococcus aureus is an important pathogenic factor of the bacterium S. aureus and also a model system for the process of membrane-induced protein oligomerisation and pore formation. It has been shown that binding to lipid membranes at neutral or basic pH requires the presence of a phosphocholine-headgroup. Thus, sphingomyelin and phosphatidylcholine may serve as interaction partners in cellular membranes. Based on earlier studies it has been suggested that rafts of sphingomyelin are particularly efficient in toxin binding. In this study we compared the oligomerisation of Hla on liposomes of various lipid compositions in order to identify the preferred interaction partners and conditions. Hla seems to have an intrinsic preference for sphingomyelin compared to phosphatidylcholine due to a higher probability of oligomerisation of membrane bound monomer. We also can show that increasing the surface density of Hla-binding sites enhances the oligomerisation efficiency. Thus, preferential binding to lipid rafts can be expected in the cellular context. On the other hand, sphingomyelin in the liquid disordered phase is a more favourable binding partner for Hla than sphingomyelin in the liquid ordered phase, which makes the membrane outside of lipid rafts the more preferred region of interaction. Thus, the partitioning of Hla is expected to strongly depend on the exact composition of raft and non-raft domains in the membrane.  相似文献   

13.
This work describes the karyotype and chromosomal location of the ribosomal DNA (rDNA) of Pecten maximus and Mimachlamys varia, two commercial scallop species from Europe. According to the chromosome centromeric index values found, the karyotype of P. maximus is composed of 1 metacentric, 2 metacentric–submetacentric, 1 telocentric–subtelocentric and 15 telocentric pairs, and that of M. varia of 4 metacentric, 2 subtelocentric–submetacentric, 9 subtelocentric, 3 subtelocentric–telocentric and 1 telocentric–subtelocentric pairs. In P. maximus, 18S-28S rDNA was located by FISH on a metacentric–submetacentric pair, and in M. varia on a subtelocentric–submetacentric pair using both silver staining and FISH. PCR amplification of the 5S rDNA unit yielded a single product of about 460 bp (P. maximus) and 450 bp (M. varia), that used as probe revealed a 5S rDNA site on a telocentric pair in P. maximus and a subtelocentric pair in M. varia. Two-color FISH or sequential silver staining of 5S rDNA-FISH-metaphases corroborated that the two gene families are located on different chromosomes in both species. A comparative analysis of the data allowed the inference of karyotypic relationships within scallops.  相似文献   

14.
15.
A thermotolerant -amylase was purified from Bacillus circulans S31 isolated from soil in Hong Kong. The purified enzyme has an M r of 64 kDa and was stable at 50°C and pH 7.0 for 30 min. Its K m for starch was 0.9 mg/ml with a V max of 0.3 mg/min. It was not activated by any metal ion although sulphydrys reagents were inhibitory.H.S. Kwan, K.H. So and K.Y. Chan are with the Department of Biology, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong S.C. Cheng is with the Department of Applied Biology and Chemical Technology, Hong Kong Polytechnic, Hung Hom, Hong Kong.  相似文献   

16.
17.
αS-Casein, the major milk protein, comprises αS1- and αS2-casein and acts as a molecular chaperone, stabilizing an array of stressed target proteins against precipitation. Here, we report that αS-casein acts in a similar manner to the unrelated small heat-shock proteins (sHsps) and clusterin in that it does not preserve the activity of stressed target enzymes. However, in contrast to sHsps and clusterin, α-casein does not bind target proteins in a state that facilitates refolding by Hsp70. αS-Casein was also separated into α- and α-casein, and the chaperone abilities of each of these proteins were assessed with amorphously aggregating and fibril-forming target proteins. Under reduction stress, all α-casein species exhibited similar chaperone ability, whereas under heat stress, α-casein was a poorer chaperone. Conversely, αS2-casein was less effective at preventing fibril formation by modified κ-casein, whereas α- and αS1-casein were comparably potent inhibitors. In the presence of added salt and heat stress, αS1-, α- and αS-casein were all significantly less effective. We conclude that αS1- and α-casein stabilise each other to facilitate optimal chaperone activity of αS-casein. This work highlights the interdependency of casein proteins for their structural stability.  相似文献   

18.
Clonal reproduction may contribute to plant evolution either by affecting population biology or by allowing partially sterile individuals (e.g., hybrids) repeated opportunities to reproduce sexually. The interplay of hybridization and clonal reproduction was first proposed by Stebbins (1950), but previously has not been tested experimentally. Alternative models for the effects of hybridization include speciation, introgression, and swamping. Salix spp. were chosen to test comparatively these hypotheses because they are easily hybridized and cloned. Over 500 separate crosses and backcrosses were made and over 6000 separate plants were measured in field experiments and statistically compared for significance to both evolutionary theory and plant breeding for biomass production. The F1 hybrids in this study always equalled, and in the case of the hybrid PR, outperformed their parents in vegetative parameters. It seems likely that even without reproducing sexually, these F1 hybrids could exist as successful individuals (sensu Stebbins 1950). However, it also seems likely that they would sexually reproduce: three of four F1 hybrids studied (RX, RP, and PR) equalled or surpassed their parents in sexual parameters when crossing with at least one other accession. Of the alternative models, experimental data suggest that introgression would be the most likely outcome of a hybridization event. The hybrid XR, however, was partially sterile and performed poorly when crossing with all other accessions in its group except S. exigua (pistillate parent). Thus, this hybrid may fit Stebbins' model of a partially sterile yet vegetatively vigorous plant that can exist as a successful individual and make some contribution to interspecific gene flow over time. This is the first experimental study to confirm the evolutionary importance of clonal reproduction coupled with hybridization. However, distinguishing any of these evolutionary pathways would be difficult in nature using morphological techniques, as interspecific hybrids tend to resemble their pistillate parents in terms of leaf shape.  相似文献   

19.
The most widely cultivated species of cotton,Gossypium hirsutum, is a disomic tetraploid (2n=4x=52). It has been proposed previously that extant A- and D-genome species are most closely related to the diploid progenitors of the tetraploid. We used fluorescent in situ hybridization (FISH) to determine the distribution of 5S and 18S-28S rDNA loci in the A-genome speciesG. herbaceum andG. arboreum, the D-genome speciesG. raimondii andG. thurberi, and the AD tetraploidG. hirsutum. High signal-to-noise, single-label FISH was used to enumerate rDNA loci, and simultaneous, dual-label FISH was used to determine the syntenic relationships of 5S rDNA loci relative to 18S–28S rDNA loci. These techniques provided greater sensitivity than our previous methods and permitted detection of six newG. hirsutum 18S–28S rDNA loci, bringing the total number of observed loci to 11. Differences in the intensity of the hybrizization signal at these loci allowed us to designate them as major, intermediate, or minor 18–28S loci. Using genomic painting with labeled A-genome DNA, five 18S–28S loci were localized to theG. hirsutum A-subgenome and six to the D-subgenome. Four of the 11 18S–28S rDNA loci inG. hirsutum could not be accounted for in its presumed diploid progenitors, as both A-genome species has three loci and both D-genome species had four.G. hirsutum has two 5S rDNA loci, both of which are syntenic to major 18S–28S rDNA loci. All four of the diploid genomes wer examined contained a single 5S locus. InG. herbaceum (A1) andG. thurberi (D1), the 5S locus is syntenic to a major 18S–28S locus, but inG. arboreum (A2) andG. raimondii (D5), the proposed D-genome progenitor ofG. hirsutum, the 5S loci are syntenic tominor and intermediate 18S–28S loci, respecitively. The multiplicity, variation in size and site number, and lack of additivity between the tetraploid species and its putative diploid ancestors indicate that the behavior of rDNA loci in cotton is nondogmatic, and considerably more complex and dynamic than previously envisioned. The relative variability of 18S–28S rDNA loci versus 5S rDNA loci suggests that the behavior of tandem repearts can differ widely. Edited by: R. Appels  相似文献   

20.

Objective

To analyze the association between drug (DAD) and alcohol (AAD) abuse and dependency and criminal and clinical background by gender of prisoners in São Paulo, Brazil.

Method

Cross-sectional study, random sample stratified by administrative district, from which prisons and prisoners were selected via random, multistage sampling. Psychiatric diagnoses were made with the CIDI 2.1. Lifetime prevalence and 95% CI were calculated and adjusted via analysis of complex samples. Multinomial logistic regression analysis was carried out with four categories of dependent variables: presence AAD; presence DAD; presence of another mental disorder; no mental disorders. For female alcohol and drug abuse and dependency (ADAD) were combined into a single category.

Results

The sample was composed by 1809 interviewed prisoners (1192 men and 617 women). Prevalence of DAD and AAD was 25.2% and 15.6%, respectively, among female prisoners, and 26.5% and 18.5% among males. Male prisoners with DAD were more likely to have a criminal record as an adolescent (OR 2.17), to be a repeat offender (OR 2.85), and to have committed a property crime (OR 2.18). Prisoners with AAD were repeat offenders (OR 2.18). Among female prisoners, ADAD was associated with repeat offenses (OR 3.39), a criminal record as an adolescent (OR 9.24), a clinical or infectious condition (OR 5.09), another health problem (OR 3.04), and violent crime (OR 2.5).

Conclusion

The study confirmed an association between drug-use disorders and the criminal and clinical background in the study population. Prisoners with such disorders were more likely to be repeat offenders and to have a criminal record as adolescents. Among female prisoners disorders were also associated with violent crime and health problems, while among males they were associated with property crime. These patterns in clinical and criminal backgrounds illustrate the need for social rehabilitation programs and specific medical treatment for prison populations.  相似文献   

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