Many secretory and several vacuolar proteins in higher plants contain hydroxylated proline residues. In many cases, hydroxyprolines in proteins are glycosylated with either arabinogalactan or ...oligoarabinose. We have previously shown that a sporamin precursor is O-glycosylated at the hydroxylated proline 36 residue with an arabinogalactan-type glycan when this protein is expressed in tobacco BY-2 cells ( Matsuoka et al., 1995 ). Taking advantage of the fact that this is the only site of proline hydroxylation and glycosylation in sporamin, we analyzed the amino acid requirement for proline hydroxylation and arabinogalactosylation. We expressed several deletion constructs and many amino acid substitution mutants in tobacco cells and analyzed glycosylation and proline hydroxylation of the expressed sporamins. Hydroxylation of a proline residue requires the five amino acid sequence AVSTG-Pro-AVSTGA-GAVPSTC-APS or acidic (where Pro is the modification site) and glycosylation of hydroxyproline (Hyp) requires the seven amino acid sequence not basic-not T-neither P, T, nor amide-Hyp-neither amide nor P-not amide-APST, although charged amino acids at the -2 position and basic amide residues at the +1 position relative to the modification site seem to inhibit the elongation of the arabinogalactan side chain. Based on the combination of these two requirements, we concluded that the sequence motif for efficient arabinogalactosylation, including the elongation of the glycan side chain, is not basic-not T-AVSG-Pro-AVST-GAVPSTC-APS.
SPCs (subtilisin-like proprotein convertases) are a family of seven structurally related serine endoproteases that are involved in the proteolytic activation of proproteins. In an effort to examine ...the substrate protein for PACE4 (paired basic amino-acid-cleaving enzyme-4), an SPC, a potent protein inhibitor of PACE4, an alpha1-antitrypsin RVRR (Arg-Val-Arg-Arg) variant, was expressed in GH4C1 cells. Ectopic expression of the RVRR variant caused accumulation of the 48 kDa protein in cells. Sequence analysis indicates that the 48 kDa protein is a putative Ca2+-binding protein, RCN-3 (reticulocalbin-3), which had previously been predicted by bioinformatic analysis of cDNA from the human hypothalamus. RCN-3 is a member of the CREC (Cab45/reticulocalbin/ERC45/calumenin) family of multiple EF-hand Ca2+-binding proteins localized to the secretory pathway. The most interesting feature of the RCN-3 sequence is the presence of five Arg-Xaa-Xaa-Arg motifs, which represents the target sequence of SPCs. Biosynthetic studies showed that RCN-3 is transiently associated with proPACE4, but not with mature PACE4. Inhibition of PACE4 maturation by a Ca2+ ionophore resulted in accumulation of the proPACE4-RCN-3 complex in cells. Furthermore, autoactivation and secretion of PACE4 was increased upon co-expression with RCN-3. Our findings suggest that selective and transient association of RCN-3 with the precursor of PACE4 plays an important role in the biosynthesis of PACE4.
This paper reports the experimental study of a 400 V class DC microgrid for an office building that has been constructed in Obihiro City, Hokkaido, Japan. The objective of this study is to develop a ...self-sustained distributed energy system by combining distribution energies, batteries, and appliances with DC power. This new DC energy system not only reduces the environment load and improves energy efficiency but also forms a community energy system that can become independent from utility grids and resistant to natural disasters. We have found that compared with AC power supply, the DC system, which uses power generated by solar panels as is, decreases CO2 emission more than 10%.
Mass tuortality atuongs seabass larvae, Lates calcarifer, reared in hatcheries in East and Bali due to viral nervous necrosis were investigated. outbreaks of the disease occured from August to ...Novetnber 1997.
The myogenic potential of bone marrow and fetal liver cells was examined using donor cells from green fluorescent protein (GFP)-gene transgenic mice transferred into chimeric mice. Lethally ...irradiated X-chromosome-linked muscular dystrophy (mdx) mice receiving bone marrow cells from the transgenic mice exhibited significant numbers of fluorescence(+) and dystrophin(+) muscle fibres. In order to compare the generating capacity of fetal liver cells with bone marrow cells in neonatal chimeras, these two cell types from the transgenic mice were injected into busulfantreated normal or mdx neonatal mice, and muscular generation in the chimeras was examined. Cardiotoxin-induced (or -uninduced, for mdx recipients) muscle regeneration in chimeras also produced fluorescence(+) muscle fibres. The muscle reconstitution efficiency of the bone marrow cells was almost equal to that of fetal liver cells. However, the myogenic cell frequency was higher in fetal livers than in bone marrow. Among the neonatal chimeras of normal recipients, several fibres expressed the fluorescence in the cardiotoxin-untreated muscle. Moreover, fluorescence(+) mononuclear cells were observed beneath the basal lamina of the cardiotoxin-untreated muscle of chimeras, a position where satellite cells are localizing. It was also found that mononuclear fluorescence(+) and desmin(+) cells were observed in the explantation cultures of untreated muscles of neonatal chimeras. The fluorescence(+) muscle fibres were generated in the second recipient mice receiving muscle single cells from the cardiotoxin-untreated neonatal chimeras. The results suggest that both bone marrow and fetal liver cells may have the potential to differentiate into muscle satellite cells and participate in muscle regeneration after muscle damage as well as in physiological muscle generation.
A new oral vaccine for Alzheimer's disease was developed using recombinant adeno-associated virus vector carrying A beta cDNA (AAV/A beta ). Oral administration of the vaccine without adjuvant ...induced the expression and secretion of A beta 1-43 or A beta 1-21 in the epithelial cell layer of the intestine in amyloid precursor protein transgenic mice. Serum antibody levels were elevated for more than six months, while T cell proliferative responses to A beta was not detected. Brain A beta burden was significantly decreased compared to the control without inflammatory changes. This oral AAV/A beta vaccine seems to be promising for prevention and treatment of Alzheimer's disease.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
We isolated a human cAMP-specific phosphodiesterase (PDE7B) cDNA from human caudate nucleus. The human PDE7B was composed of 450 amino acid residues with a molecular mass of 51,835 Da. The deduced ...amino acid sequence of human PDE7B was 64.1% identical to that of human PDE7A (67.1% identity in the catalytic region). Northern blot analysis demonstrated that PDE7B transcripts were abundantly expressed in the putamen, caudate nucleus, and heart followed by skeletal muscle, pancreas, and occipital pole. Recombinant PDE7B expressed in transfected COS-7 cells had a low cAMP Km value of 0.13 μM, which is similar to the Km value of recombinant human PDE7A expressed in transfected COS-7 cells. Interestingly, the relative Vmax value of recombinant PDE7B was half to one-third of recombinant PDE7A. The PDE7B activity was inhibited by dipyridamole and SCH51866, with IC50 values of 1.1 μM and 1.5 μM, respectively. Thus, the PDE7B exhibited unique tissue distribution in humans and kinetic profiles. Human PDE7B showed the lowest Km values compared to the other cAMP-hydrolyzing PDEs which have been reported to be expressed in the brain. Therefore, human PDE7B may be involved in the control of cAMP-mediated neural activity and cAMP metabolism in the brain.
The C-type natriuretic peptide/natriuretic peptide receptor-B/cGMP pathway plays an important role in the regulation of endochondral ossification. In chondrocytes, the physiological effect of cGMP is ...mediated primarily by the activation of cGMP-dependent protein kinase II (cGK-II). In this study, we investigated the transcriptional regulation of cGK-II in chondrocytes. The expression pattern of cGK-II transcripts was examined during chondrogenic differentiation of ATDC5 cells. cGK-II mRNA was not detectable in undifferentiated cells, but increased dramatically prior to differentiation to the hypertrophic stage. To analyze the transcriptional regulation of cGK-II, the 5′-flanking region of the mouse cGK-II gene was isolated and characterized. The promoter activity of the cGK-II gene decreased markedly following deletion and mutagenesis of the putative Nkx-binding site between nucleotide positions −292 and −286. These results suggest that the homeobox gene Nkx family is critical for the transcriptional regulation of cGK-II during chondrogenesis.
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