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1.
The plant-specific insert is an approximately 100-residue domain found exclusively within the C-terminal lobe of some plant aspartic proteases. Structurally, this domain is a member of the saposin-like protein family, and is involved in plant pathogen defense as well as vacuolar targeting of the parent protease molecule. Similar to other members of the saposin-like protein family, most notably saposins A and C, the recently resolved crystal structure of potato (Solanum tuberosum) plant-specific insert has been shown to exist in a substrate-bound open conformation in which the plant-specific insert oligomerizes to form homodimers. In addition to the open structure, a closed conformation also exists having the classic saposin fold of the saposin-like protein family as observed in the crystal structure of barley (Hordeum vulgare L.) plant-specific insert. In the present study, the mechanisms of tertiary and quaternary conformation changes of potato plant-specific insert were investigated in silico as a function of pH. Umbrella sampling and determination of the free energy change of dissociation of the plant-specific insert homodimer revealed that increasing the pH of the system to near physiological levels reduced the free energy barrier to dissociation. Furthermore, principal component analysis was used to characterize conformational changes at both acidic and neutral pH. The results indicated that the plant-specific insert may adopt a tertiary structure similar to the characteristic saposin fold and suggest a potential new structural motif among saposin-like proteins. To our knowledge, this acidified PSI structure presents the first example of an alternative saposin-fold motif for any member of the large and diverse SAPLIP family. 相似文献
2.
The aim of our work was to study the mechanism of action of aldosterone and antialdosterone compounds on Na+ and K+ fluxes in vascular smooth muscle. In the long term, regulation of salt metabolism depends on aldosterone effects on Na+, K+, H+ and H2O transport by the renal tubules. Furthermore, it has been shown that aldosterone modifies several epithelial transports, inducing a positive sodium balance. The chronic in vivo administration of aldosterone modifies transmembrane ionic fluxes in vascular smooth muscle. Garwitz and Jones suggested that aldosterone may enhance net Na+ transport through the stimulation of the sodium pump. The results obtained in our laboratory indicate that aldosterone has a direct stimulatory action on ouabain-dependent and on ouabain-independent Na efflux. Furthermore, the mineralocorticoid enhances passive K permeability, as well as the Na pump dependent K influx. Both effects are blocked by antimineralocorticoid compounds. Recent experiments have shown that vasopressin potentiates some of the in vivo effects of aldosterone. 相似文献
3.
The three classes of hydrogenases from sulfate-reducing bacteria of the genus Desulfovibrio 总被引:12,自引:0,他引:12
G Fauque H D Peck J J Moura B H Huynh Y Berlier D V DerVartanian M Teixeira A E Przybyla P A Lespinat I Moura 《FEMS microbiology reviews》1988,4(4):299-344
Three types of hydrogenases have been isolated from the sulfate-reducing bacteria of the genus Desulfovibrio. They differ in their subunit and metal compositions, physico-chemical characteristics, amino acid sequences, immunological reactivities, gene structures and their catalytic properties. Broadly, the hydrogenases can be considered as 'iron only' hydrogenases and nickel-containing hydrogenases. The iron-sulfur-containing hydrogenase ([Fe] hydrogenase) contains two ferredoxin-type (4Fe-4S) clusters and an atypical iron-sulfur center believed to be involved in the activation of H2. The [Fe] hydrogenase has the highest specific activity in the evolution and consumption of hydrogen and in the proton-deuterium exchange reaction and this enzyme is the most sensitive to CO and NO2-. It is not present in all species of Desulfovibrio. The nickel-(iron-sulfur)-containing hydrogenases [( NiFe] hydrogenases) possess two (4Fe-4S) centers and one (3Fe-xS) cluster in addition to nickel and have been found in all species of Desulfovibrio so far investigated. The redox active nickel is ligated by at least two cysteinyl thiolate residues and the [NiFe] hydrogenases are particularly resistant to inhibitors such as CO and NO2-. The genes encoding the large and small subunits of a periplasmic and a membrane-bound species of the [NiFe] hydrogenase have been cloned in Escherichia (E.) coli and sequenced. Their derived amino acid sequences exhibit a high degree of homology (70%); however, they show no obvious metal-binding sites or homology with the derived amino acid sequence of the [Fe] hydrogenase. The third class is represented by the nickel-(iron-sulfur)-selenium-containing hydrogenases [( NiFe-Se] hydrogenases) which contain nickel and selenium in equimolecular amounts plus (4Fe-4S) centers and are only found in some species of Desulfovibrio. The genes encoding the large and small subunits of the periplasmic hydrogenase from Desulfovibrio (D.) baculatus (DSM 1743) have been cloned in E. coli and sequenced. The derived amino acid sequence exhibits homology (40%) with the sequence of the [NiFe] hydrogenase and the carboxy-terminus of the gene for the large subunit contains a codon (TGA) for selenocysteine in a position homologous to a codon (TGC) for cysteine in the large subunit of the [NiFe] hydrogenase. EXAFS and EPR studies with the 77Se-enriched D. baculatus hydrogenase indicate that selenium is a ligand to nickel and suggest that the redox active nickel is ligated by at least two cysteinyl thiolate and one selenocysteine selenolate residues.(ABSTRACT TRUNCATED AT 400 WORDS) 相似文献
4.
EPR-detectable redox centers of the periplasmic hydrogenase from Desulfovibrio vulgaris 总被引:1,自引:0,他引:1
D S Patil J J Moura S H He M Teixeira B C Prickril D V DerVartanian H D Peck J LeGall B H Huynh 《The Journal of biological chemistry》1988,263(35):18732-18738
The periplasmic hydrogenase of Desulfovibrio vulgaris (Hildenbourough NCIB 8303) belongs to the category of [Fe] hydrogenase which contains only iron-sulfur clusters as its prosthetic groups. Amino acid analyses were performed on the purified D. vulgaris hydrogenase. The amino acid composition obtained compared very well with the result derived from the nucleotide sequence of the structural gene (Voordouw, G., Brenner, S. (1985) Eur. J. Biochem. 148, 515-520). Detailed EPR reductive titration studies on the D. vulgaris hydrogenase were performed to characterize the metal centers in this hydrogenase. In addition to the three previously observed EPR signals (namely, the "isotropic" 2.02 signal, the rhombic 2.10 signal, and the complex signal of the reduced enzyme), a rhombic signal with resonances at the g-values of 2.06, 1.96, and 1.89 (the rhombic 2.06 signal) was detected when the samples were poised at potentials between 0 and -250 mV (with respect to normal hydrogen electrode). The midpoint redox potentials for each of the four EPR-active species were determined, and the characteristics of each EPR signal are described. Both the rhombic 2.10 and 2.06 signals exhibit spectral properties that are distinct from a ferredoxin-type [4Fe-4S] cluster and are proposed to originate from the same H2-binding center but in two different conformations. The complex signal of the reduced hydrogenase has been shown to represent two spin-spin interacting ferredoxin-type [4Fe-4S]1+ clusters (Grande, H. J., Dunham, W. R., Averill, B., Van Dijk, C., and Sands, R. H. (1983) Eur. J. Biochem. 136, 201-207). The titration data indicated a strong cooperative effect between these two clusters during their reduction. In an effort to accurately estimate the number of iron atoms/molecule of hydrogenase, plasma emission and chemical methods were used to determine the iron contents in the samples; and four different methods, including amino acid analysis, were used for protein determination. The resulting iron stoichiometries were found to be method-dependent and vary over a wide range (+/- 20%). The uncertainties involved in the determination of iron stoichiometry are discussed. 相似文献
5.
A new experimental model, the latissimus dorsi flap of the rabbit, was studied. This was found to be a relatively inexpensive research model. Its use is advocated for composite tissue transfer as transposition, island, or free myocutaneous flaps. 相似文献
6.
M Teixeira I Moura A V Xavier J J Moura J LeGall D V DerVartanian H D Peck B H Huynh 《The Journal of biological chemistry》1989,264(28):16435-16450
The hydrogenase (EC 1.2.2.1) of Desulfovibrio gigas is a complex enzyme containing one nickel center, one [3Fe-4S] and two [4Fe-4S] clusters. Redox intermediates of this enzyme were generated under hydrogen (the natural substrate) using a redox-titration technique and were studied by EPR and M?ssbauer spectroscopy. In the oxidized states, the two [4Fe-4S]2+ clusters exhibit a broad quadrupole doublet with parameters (apparent delta EQ = 1.10 mm/s and delta = 0.35 mm/s) typical for this type of cluster. Upon reduction, the two [4Fe-4S]1+ clusters are spectroscopically distinguishable, allowing the determination of their midpoint redox potentials. The cluster with higher midpoint potential (-290 +/- 20 mV) was labeled Fe-S center I and the other with lower potential (-340 +/- 20 mV), Fe-S center II. Both reduced clusters show atypical magnetic hyperfine coupling constants, suggesting structural differences from the clusters of bacterial ferredoxins. Also, an unusually broad EPR signal, labeled Fe-S signal B', extending from approximately 150 to approximately 450 mT was observed concomitantly with the reduction of the [4Fe-4S] clusters. The following two EPR signals observed at the weak-field region were tentatively attributed to the reduced [3Fe-4S] cluster: (i) a signal with crossover point at g approximately 12, labeled the g = 12 signal, and (ii) a broad signal at the very weak-field region (approximately 3 mT), labeled the Fe-S signal B. The midpoint redox potential associated with the appearance of the g = 12 signal was determined to be -70 +/- 10 mV. At potentials below -250 mV, the g = 12 signal began to decrease in intensity, and simultaneously, the Fe-S signal B appeared. The transformation of the g = 12 signal into the Fe-S signal B was found to parallel the reduction of the two [4Fe-4S] clusters indicating that the [3Fe-4S]o cluster is sensitive to the redox state of the [4Fe-4S] clusters. Detailed redox profiles for the previously reported Ni-signal C and the g = 2.21 signal were obtained in this study, and evidence was found to indicate that these two signals represent two different oxidation states of the enzyme. Finally, the mechanistic implications of our results are discussed. 相似文献
7.
EPR studies with 77Se-enriched (NiFeSe) hydrogenase of Desulfovibrio baculatus. Evidence for a selenium ligand to the active site nickel 总被引:5,自引:0,他引:5
S H He M Teixeira J LeGall D S Patil I Moura J J Moura D V DerVartanian B H Huynh H D Peck 《The Journal of biological chemistry》1989,264(5):2678-2682
The periplasmic hydrogenase containing equivalent amounts of nickel and selenium plus non-heme iron [NiFeSe) hydrogenase) has been purified from cells of the sulfate reducing bacterium Desulfovibrio baculatus (DSM 1748) grown on a lactate/sulfate medium containing natural Se isotopes and the nuclear isotope, 77Se. Both the 77Se-enriched and unenriched hydrogenases were shown to be free of other hydrogenases and characterized with regard to their Se contents. EPR studies of the reduced nickel signal generated by redox titrations of the enriched and unenriched (NiFeSe) hydrogenases demonstrated that the gx = 2.23 and gy = 2.17 resonances are appreciably broadened by the spin of the 77Se nucleus (I = 1/2). This observation demonstrates unambiguously that the unpaired electron is shared by the Ni and Se atoms and that Se serves as a ligand to the nickel redox center of the (NiFeSe) hydrogenase. 相似文献
8.
de Sampaio TC Melo RB Moura TF Michel S Barreiros S 《Biotechnology and bioengineering》1996,50(3):257-264
We studied a model transesterification reaction catalyzed by subtilisin Carlsberg suspended in toluene, n-hexane, diisopropyl ether, and mixtures of these solvents. To account for solvent effects due to differences in water partitioning between the enzyme and the bulk solvents, we measured water sorption isotherms for the enzyme in each solvent. We measured catalytic activity as a function of enzyme hydration and obtained bell-shaped curves with maxima at the same enzyme hydration in all the solvents. However, the activity maxima were different in all the media, being the lowest in toluene. Differences in the partitioning of substrates and product between the bulk solvent phase and the enzyme active site were accounted for but could not explain the lower catalytic activity observed in toluene. The fact that toluene is very similar to one of the substrates suggested the possibility of competitive inhibition by this solvent. We derived a model allowing for differences in solvation of the substrates, by using thermodynamic activities instead of concentrations, as well as for competitive inhibition by toluene. The model fit the experimental data well, confirming that toluene had a direct adverse effect on the catalytic activity of the enzyme. (c) 1996 John Wiley & Sons, Inc. 相似文献
9.
Nielsen J; Peixoto AA; Piccin A; Costa R; Kyriacou CP; Chalmers D 《Molecular biology and evolution》1994,11(6):839-853
The region of the clock gene period (per) that encodes a repetitive tract
of threonine-glycine (Thr-Gly) pairs has been compared between Dipteran
species both within and outside the Drosophilidae. All the non-
Drosophilidae sequences in this region are short and present a remarkably
stable picture compared to the Drosophilidae, in which the region is much
larger and extremely variable, both in size and composition. The
accelerated evolution in the repetitive region of the Drosophilidae appears
to be mainly due to an expansion of two ancestral repeats, one encoding a
Thr-Gly dipeptide and the other a pentapeptide rich in serine, glycine, and
asparagine or threonine. In some drosophilids the expansion involves a
duplication of the pentapeptide sequence, but in Drosophila pseudoobscura
both the dipeptide and the pentapeptide repeats are present in larger
numbers. In the nondrosophilids, however, the pentapeptide sequence is
represented by one copy and the dipeptide by two copies. These observations
fulfill some of the predictions of recent theoretical models that have
simulated the evolution of repetitive sequences.
相似文献
10.
Prof. Christchellyn Klegin Prof. Dr. Eduardo Miranda Ethur Bárbara Buhl Ana Caroline Giacomin Prof. Dr. Juçara Bordin Prof. Dr. Neusa Fernandes de Moura Matheus Henrique Oliveira de Sousa 《化学与生物多样性》2023,20(6):e202300253
The present study aimed to examine the phenolic content and evaluate the antimicrobial and antioxidant potential of ethanol extracts from the moss species Phyllogonium viride Brid. on the pathogenic bacteria Salmonella enterica serovar enteritidis, Staphylococcus aureus, Listeria monocytogenes and Escherichia coli, and the pathogenic fungi Candida albicans and Cryptococcus neoformans. The antimicrobial activity was determined from Minimum Inhibitory Concentration (MIC) Minimum Bactericidal Concentration (MBC) and Minimum Fungicidal Concentration (MFC). Antioxidant activity was determined by the DPPH method. Folin-Denis reagent was used for the content of total phenolics and flavonoids and HPLC-DAD for identification of phenolic compounds. The results showed that bacteriostatic and bactericidal activities occurred at concentrations ranging from 9.76 μg/mL–78.13 μg/mL among all evaluated microorganisms. These values, considering the criteria used, suggest the P. viride extract as a potent antimicrobial. For antioxidant activity, P. viride extract was considered weak. Analysis of the phenolic content showed a wide range of compounds, with Kaempferol (0.41 mg/g) being the major compound, followed by t-cinnamic acid and caffeic acid (0.17 mg/g). Although P. viride is a species of moss not yet referenced in scientific publications of biotechnological interest, it has shown promising potential for further studies and possible application as an antimicrobial of natural origin. 相似文献