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11.
Nanna Alho Tuula Klaavuniemi Jari Ylänne Perttu Permi Sampo Mattila 《Biomolecular NMR assignments》2007,1(1):85-87
Earlier reports have shown that ALP has an internal interaction site. We were able to stablize the structure of this unfolded
part to a great extent by aspartic acid, which allowed the backbone assignment. No secondary structure of the polypeptide
was observed. 相似文献
12.
The plant plasma membrane (PM) H+-ATPase is an essential enzyme controlling plant growth and development. It is an important factor in response to abiotic and biotic stresses and is subject to tight regulation. We are in demand for new sustainable natural growth regulators and as a key enzyme for regulation of transport into the plant cell the PM H+-ATPase is a potential target for these. In this review, we have evaluated the known non-protein natural compounds with regulatory effects on the PM H+-ATPase, focusing on their mechanism of action and their potential as biologicals/growth regulators in plant production of future sustainable agriculture. 相似文献
13.
The amino acid sequence of human myoglobin (Mb) is similar to other mammalian Mb except for a unique cysteine residue at position 110 (Cys(110)). Anaerobic treatment of ferrous forms of wild-type human Mb, the C110A variant of human Mb or horse heart Mb, with either authentic NO or chemically derived NO in vitro yields heme-NO complexes as detected by electron paramagnetic resonance spectroscopy (EPR). By contrast, no EPR-detectable heme-NO complex was observed from the aerobic reactions of NO and either the ferric or oxy-Mb forms of wild-type human or horse heart myoglobins. Mass analyses of wild-type human Mb treated aerobically with NO indicated a mass increase of approximately 30 atomic mass units (i.e., NO/Mb = 1 mol/mol). Mass analyses of the corresponding apoprotein after heme removal showed that NO was associated with the apoprotein fraction. New electronic maxima were detected at A(333 nm) (epsilon = 3665 +/- 90 mol(-)(1) cm(-)(1); mean +/- S.D.) and A(545 nm) (epsilon = 44 +/- 3 mol(-)(1) cm(-)(1)) in solutions of S-nitrosated wild-type human Mb (similar to S-nitrosoglutathione). Importantly, the sulfhydryl S-H stretch vibration for Cys(110) measured by Fourier transform infrared (nu approximately 2552 cm(-)(1)) was absent for both holo- and apo- forms of the wild-type human protein after aerobic treatment of the protein with NO. Together, these data indicate that the reaction of wild-type human Mb and NO yields either heme-NO or a novel S-nitrosated protein dependent on the oxidation state of the heme iron and the presence or absence of dioxygen. 相似文献
14.
RhoA exerts a permissive effect on volume-regulated anion channels in vascular endothelial cells 总被引:2,自引:0,他引:2
Carton I Trouet D Hermans D Barth H Aktories K Droogmans G Jorgensen NK Hoffmann EK Nilius B Eggermont J 《American journal of physiology. Cell physiology》2002,283(1):C115-C125
Cell swelling triggers in most cell typesan outwardly rectifying anion current,ICl,swell, via volume-regulated anion channels (VRACs). We have previously demonstrated in calf pulmonary artery endothelial (CPAE) cells that inhibition of the Rho/Rho kinase/myosin light chain phosphorylation pathway reduces the swelling-dependent activation of ICl,swell. However, theseexperiments did not allow us to discriminate between a direct activatorrole or a permissive effect. We now show that the Rho pathway did notaffect VRAC activity if this pathway was activated by transfecting CPAEcells with constitutively active isoforms of G (a Rho activatingheterotrimeric G protein subunit), Rho, or Rho kinase. Furthermore,biochemical and morphological analysis failed to demonstrate activationof the Rho pathway during hypotonic cell swelling. Finally,manipulating the Rho pathway with either guanosine5'-O-(3-thiotriphosphate) or C3 exoenzyme had no effect onVRACs in caveolin-1-expressing Caco-2 cells. We conclude that the Rhopathway exerts a permissive effect on VRACs in CPAE cells, i.e.,swelling-induced opening of VRACs requires a functional Rho pathway,but not an activation of the Rho pathway. 相似文献
15.
Andriambeloson E Witting PK 《Redox report : communications in free radical research》2002,7(3):131-136
The detailed chemistry of nitric oxide (*NO) and regulation of this potent signal molecule through interactions with cellular components are complex and not clearly understood. In the vasculature, *NO plays a crucial role in vessel dilation by activating soluble guanylyl cyclase (sGC) in vascular smooth muscle cells (VSMC). *NO is responsible for maintaining coronary blood flow and normal cardiac function. However, *NO is a highly reactive molecule and this reactivity toward a range of alternate substrates may interfere with the activation of its preferred molecular target within VSMC. Interestingly, marked changes to *NO homeostasis are linked to disease progression. Thus, the physiological concentration of *NO is carefully regulated. Myoglobin is a haem-containing protein that is present in relatively high concentration in cardiac and skeletal muscle. Recently, the presence of myoglobin has been confirmed in human smooth muscle. The role of intracellular myoglobin is generally accepted as that of a passive di-oxygen storage protein. However, oxygenated myoglobin readily reacts with *NO to yield higher order N-oxides such as nitrate, while both the ferrous and ferric forms of the protein form a stable complex with *NO. Together, these two reactions effectively eliminate *NO on the physiological time-scale and strongly support the idea that myoglobin plays a role in maintaining *NO homeostasis in tissues that contain the protein. Interestingly, human myoglobin contains a sulfhydryl group and forms an S-nitroso-adduct similar to haemoglobin. In this article we discuss the potential for human myoglobin to actively participate in the regulation of *NO by three distinct mechanisms, namely oxidation, ligand binding, and through formation of biologically active S-nitroso-myoglobin. 相似文献
16.
Panagotopulos SE Witting SR Horace EM Nicholas Maiorano J Sean Davidson W 《Protein expression and purification》2002,25(2):353-361
Plasma levels of apolipoprotein A-I (apoA-I) are correlated with reduced incidence of heart disease due to the critical role of this protein in reverse cholesterol transport. Because of its diversity of function and poorly understood structure, much research has sought to understand how the structure of apoA-I facilitates its function. A popular approach has been the use of site-directed mutagenesis followed by structural and functional studies. There are a wide variety of expression systems available to produce these mutant proteins including eukaryotic cell lines and prokaryotic cells such as Escherichia coli. Expression in a bacterial system is generally favorable because it can produce large amounts of pure protein quickly and economically through the use of affinity tags on the expressed protein. Unfortunately, many of these systems are not ideal for the production of apolipoproteins because, in many cases, the proteolytic digestion required to remove the affinity tag also cleaves the target protein. Here we describe a method that produces large amounts of recombinant protein that is easily purified using a histidine (His) affinity tag that is cleaved with IgA protease from Neisseria gonorrhoeae. This enzyme does not cleave the wild type apoA-I sequence, leaving intact, mature apoA-I (containing a Thr-Pro- on the N-terminus). We show that this recombinant protein is similar to wild type protein in structure and function using circular dichroism analysis, lipid clearance assays, recombinant particle formation and cholesterol efflux assays. This system is particularly useful for the bacterial production of apolipoproteins because of the extreme specificity of IgA protease for its target cleavage site. 相似文献
17.
Christensen NK Petersen M Vester B Nielsen P 《Nucleosides, nucleotides & nucleic acids》2003,22(5-8):1143-1145
Two pyrimidine alpha-LNA nucleoside monomers have been synthesised and incorporated into alpha-configured oligonucleotides. A fully modified mixed alpha-LNA sequence displays unprecedented parallel stranded hybridisation with complementary RNA and a remarkable selectivity for RNA over DNA. Modelling shows alpha-LNA:RNA to form an extended duplex with a very broad major groove. 相似文献
18.
Albaek N Ravn J Freitag M Thomasen H Christensen NK Petersen M Nielsen P 《Nucleosides, nucleotides & nucleic acids》2003,22(5-8):723-725
Ring-closing metathesis (RCM) is applied as a new and powerful technology in the construction of nucleoside analogues that are conformationally restricted in S-type conformations due to additional 3',4'- and/or 3',5'-linkages. 相似文献
19.
Sørensen NM Kobaek-Larsen M Bonne A van Zutphen B Fenger C Kristiansen K Ritskes-Hoitinga M 《Comparative medicine》2003,53(6):633-638
The aim of the study reported here was to investigate whether the azoxymethane (AOM)-induced colon cancer rat model mimics the human situation with regard to microsatellite stability, changes in expression of beta-catenin, and/or changes in the sequence of the proto-oncogene Ki-ras. Colon cancer was induced by administration of four weekly doses of AOM (15 mg/kg of body weight per week) separated by a one-week break between the second and third injections. As the histopathologic characteristics of this model resemble those of the human counterpart, further characterization of the genetic changes was undertaken. The animals were euthanized 28 to 29 weeks after the first AOM injection, and tumor specimens were taken for histologic and DNA analyses. Since microsatellite variation was found in only a few (< 2%) specimens, the model can be considered as having stable microsatellites. This result is in accordance with those of similar studies in other rat models and with most human colorectal cancers. Immunohistochemical analyses of beta-catenin did not reveal loss of gene activity, nor did the sequencing of Ki-ras reveal mutations. These results are in contrast to most findings in comparable rat studies. The deviations may be due to differences in exposure to the carcinogen or difference in strain and/or age. The lack of beta-catenin and Ki-ras alterations in this colon cancer model is unlike human sporadic colorectal cancers where these genetic changes are common findings. 相似文献
20.
The human myoglobin (Mb) sequence is similar to other mammalian Mb sequences, except for a unique cysteine at position 110. Reaction of wild-type recombinant human Mb, the C110A variant of human Mb, or horse heart Mb with H(2)O(2) (protein/H(2)O(2) = 1:1.2 mol/mol) resulted in formation of tryptophan peroxyl (Trp-OO( small middle dot)) and tyrosine phenoxyl radicals as detected by EPR spectroscopy at 77 K. For wild-type human Mb, a second radical (g approximately 2. 036) was detected after decay of Trp-OO( small middle dot) that was not observed for the C110A variant or horse heart Mb. When the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was included in the reaction mixture at protein/DMPO ratios =1:10 mol/mol, a DMPO adduct exhibiting broad absorptions was detected. Hyperfine couplings of this radical indicated a DMPO-thiyl radical. Incubation of wild-type human Mb with thiol-blocking reagents prior to reaction with peroxide inhibited DMPO adduct formation, whereas at protein/DMPO ratios >/=1:25 mol/mol, DMPO-tyrosyl radical adducts were detected. Mass spectrometry of wild-type human Mb following reaction with H(2)O(2) demonstrated the formation of a homodimer (mass of 34,107 +/- 5 atomic mass units) sensitive to reducing conditions. The human Mb C110A variant afforded no dimer under identical conditions. Together, these data indicate that reaction of wild-type human Mb and H(2)O(2) differs from the corresponding reaction of other myoglobin species by formation of thiyl radicals that lead to a homodimer through intermolecular disulfide bond formation. 相似文献