共查询到20条相似文献,搜索用时 267 毫秒
1.
Brandsch M 《Amino acids》2006,31(2):119-136
Summary. Membrane transport of L-proline has received considerable attention in basic and pharmaceutical research recently. Of the
most recently cloned members of the solute carrier family, two are “proline transporters”. The amino acid transporter PAT1,
expressed in intestine, kidney, brain and other organs, mediates the uptake of proline and derivatives in a pH gradient-dependent
manner. The Na+-dependent proline transporter SIT1, cloned in 2005, exhibits the properties of the long-sought classical IMINO system. Proline-containing
peptides are of interest for several reasons. Many biologically important peptide sequences contain highly conserved proline
residues. Xaa-Pro peptides are very often resistant to enzymatic hydrolysis and display, in contrast to Pro-Xaa peptides,
a high affinity to the H+/peptide cotransporter PEPT1 which is expressed in intestinal, renal, lung and biliary duct epithelial cells. Furthermore,
several orally available drugs are recognized by PEPT1 as Xaa-Pro analogues due to their sterical resemblance to small peptides. 相似文献
2.
The proton-coupled oligopeptide transporter PEPT2 (or SLC15A2 ) is the major protein involved in the reclamation of peptide-bound amino acids and peptide-like drugs in kidney. PEPT2 is also important in effluxing peptides and peptidomimetics from CSF at the choroid plexus, thereby limiting their exposure in brain. In this study, we report a neuroprotective role for PEPT2 in modulating the toxicity of a heme precursor, 5-aminolevulinic acid (5-ALA). Our findings demonstrate that in PEPT2-deficient mice, 5-ALA administration results in reduced survivability, a worsening of neuromuscular dysfunction, and CSF concentrations of substrate that are 8–30 times higher than that in wild-type control animals. The ability of PEPT2 to limit 5-ALA exposure in CSF suggests that it may also have relevance as a secondary genetic modifier of conditions (such as acute hepatic porphyrias and lead poisoning) in which 5-ALA metabolism is altered and in which 5-ALA toxicity is important. 相似文献
3.
Novotny A Xiang J Stummer W Teuscher NS Smith DE Keep RF 《Journal of neurochemistry》2000,75(1):321-328
5-Aminolevulinic acid (5-ALA) is a precursor of porphyrins and heme that has been implicated in the neuropsychiatric symptoms associated with porphyrias. It is also being used clinically to delineate malignant gliomas. The blood-CSF barrier may be an important interface for 5-ALA transport between blood and brain as in vivo studies have indicated 5-ALA is taken up by the choroid plexuses whereas the normal blood-brain barrier appears to be relatively impermeable. This study examines the mechanisms of 5-[(3)H]ALA uptake into isolated rat lateral ventricle choroid plexuses. Results suggest that there are two uptake mechanisms. The first was a Na(+)-independent uptake system that was pH dependent (being stimulated at low pH). Uptake was inhibited by the dipeptide Gly-Gly and by cefadroxil, an alpha-amino-containing cephalosporin. These properties are the same as the proton-dependent peptide transporters PEPT1 and PEPT2, which have recently been shown to transport 5-ALA in frog oocyte expression experiments. Choroid plexus uptake was not inhibited by captopril, a PEPT1 inhibitor, suggesting PEPT2-mediated uptake. The presence of PEPT2 and absence of PEPT1 in the choroid plexus were confirmed by western blotting. The second potential mechanism was both Na(+) and HCO(3)(-) dependent and appears to be an organic anion transporter, although it is possible that removal of Na(+) and HCO(3)(-) may indirectly affect PEPT2 by affecting intracellular pH. The presence of PEPT2 and a putative Na(+)/HCO(3)(-)-dependent organic anion transporter is important not only for an understanding of 5-ALA movement between blood and brain but also because these transporters may affect the distribution of a number of drugs between blood and CSF. 相似文献
4.
Shimizu Y Masuda S Nishihara K Ji L Okuda M Inui K 《American journal of physiology. Gastrointestinal and liver physiology》2005,288(4):G664-G670
In chronic renal failure (CRF), dietary protein is one of the factors that deteriorates residual renal functions. Numerous studies have indicated that the products of protein digestion are mainly absorbed as small peptides. However, how small peptides are absorbed in CRF remains poorly understood. H(+)-coupled peptide transporter (PEPT1/SLC15A1) plays an important role in the absorption of small peptides and peptide-like drugs in the small intestine. Because dietary protein intake is one of the risk factors for renal failure, the alteration of intestinal PEPT1 might have implications in the progression of renal disease as well as the pharmacokinetics of peptide-like drugs. In this study, we examined the alteration of intestinal PEPT1 in 5/6 nephrectomized (5/6 NR) rats, extensively used as a model of chronic renal failure. Absorption of [(14)C]glycylsarcosine and ceftibuten was significantly increased in 5/6 NR rats compared with sham-operated rats, without a change in intestinal protease activity. Western blot analysis indicated that the amount of intestinal PEPT1 protein in 5/6 NR rats was increased mainly at the upper region. On the other hand, the amount of intestinal PEPT1 mRNA was not significantly different from that of sham-operated rats. These findings indicate that the increase in absorption of small peptides and peptide-like drugs, caused by the upregulation of intestinal PEPT1 protein, might contribute to the progression of renal failure as well as the alteration of drug pharmacokinetics. 相似文献
5.
6.
Mikihisa Takano Taishi HoriuchiYoshihiro Sasaki Yuki KatoJunya Nagai Ryoko Yumoto 《Life sciences》2013
Aims
The purpose of this study was to clarify the expression and function of peptide transporter 2 (PEPT2) in primary cultured alveolar type II epithelial cells and in transdifferentiated type I-like cells.Main methods
Real-time PCR analysis, uptake study of [3H]Gly-Sar, and immunostaining were performed in alveolar epithelial cells.Key findings
The expression of PEPT2 mRNA in type II cells isolated from rat lungs was highest at day 0, and decreased rapidly during culture of the cells. In accordance with this change, PEPT2 activity estimated as cefadroxil-sensitive [3H]Gly-Sar uptake also decreased along with transdifferentiation. The expression of PEPT2 protein in type II cells was confirmed by immunostaining and Western blot analysis. The uptake of [3H]Gly-Sar in type II cells was time- and pH-dependent. In contrast, minimal time-dependence and no pH-dependence of [3H]Gly-Sar uptake were observed in type I-like cells. The maximal [3H]Gly-Sar uptake was observed at pH 6.0, and the uptake decreased at higher pHs in type II cells. The uptake of [3H]Gly-Sar in type II cells was inhibited by cefadroxil in a concentration-dependent manner, the IC50 value being 4.3 μM. On the other hand, no significant inhibition by cefadroxil was observed in type I-like cells. In addition, [3H]Gly-Sar uptake in type II cells was saturable, the Km value being 72.0 μM.Significance
PEPT2 is functionally expressed in alveolar type II epithelial cells, but the expression decreases along with transdifferentiation, and PEPT2 would be almost completely lost in type I cells. 相似文献7.
Pan X Terada T Irie M Saito H Inui K 《American journal of physiology. Gastrointestinal and liver physiology》2002,283(1):G57-G64
In mammals, most physiological, biochemical, and behavioral processes show a circadian rhythm. In the present study, we examined the diurnal rhythm of the H+-peptide cotransporter (PEPT1), which transports small peptides and peptide-like drugs in the small intestine and kidney, using rats maintained in a 12-h photoperiod with free access to chow. The transport of [14C]glycylsarcosine (Gly-Sar), a typical substrate for PEPT1 by in situ intestinal loop and everted intestine, was greater in the dark phase than the light phase. PEPT1 protein and mRNA levels varied significantly, with a maximum at 2000 and minimum at 800. Similar functional and expressional diurnal variations were observed in the intestinal Na+-glucose cotransporter (SGLT1). In contrast, renal PEPT1 and SGLT1 showed little diurnal rhythmicity in protein and mRNA expression. These findings indicate that the intestinal PEPT1 undergoes diurnal regulation in its activity and expression, and this could affect the intestinal absorption of dietary protein. 相似文献
8.
Summary. Taurine has been thought to function as a regulator of neuronal activity, neuromodulator and osmoregulator. Moreover, it is
essential for the development and survival of neural cells and protects them under cell-damaging conditions. Taurine is also
involved in many vital functions regulated by the brain stem, including cardiovascular control and arterial blood pressure.
The release of taurine has been studied both in vivo and in vitro in higher brain areas, whereas the mechanisms of release
have not been systematically characterized in the brain stem. The properties of release of preloaded [3H]taurine were now characterized in slices prepared from the mouse brain stem from developing (7-day-old) and young adult
(3-month-old) mice, using a superfusion system. In general, taurine release was found to be similar to that in other brain
areas, consisting of both Ca2+-dependent and Ca2+-independent components. Moreover, the release was mediated by Na+-, Cl−-dependent transporters operating outwards, as both Na+-free and Cl− -free conditions greatly enhanced it. Cl− channel antagonists and a Cl− transport inhibitor reduced the release at both ages, indicating that a part of the release occurs through ion channels.
Protein kinases appeared not to be involved in taurine release in the brain stem, since substances affecting the activity
of protein kinase C or tyrosine kinase had no significant effects. The release was modulated by cAMP second messenger systems
and phospholipases at both ages. Furthermore, the metabotropic glutamate receptor agonists likewise suppressed the K+-stimulated release at both ages. In the immature brain stem, the ionotropic glutamate receptor agonists N-methyl-D-aspartate
(NMDA) and 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) potentiated taurine release in a receptor-mediated manner.
This could constitute an important mechanism against excitotoxicity, protecting the brain stem under cell-damaging conditions. 相似文献
9.
Döring F Schmitt R Bernhardt WM Klapper M Bachmann S Daniel H Groneberg DA 《Biological chemistry》2005,386(8):785-790
The kidney is a target organ for thyroid hormone action and a variety of renal transport processes are altered in response to impaired thyroid functions. To investigate the effect of thyroid hormone on the expression of the renal proximal tubular high-affinity-type H(+)-peptide cotransporter (PEPT2) in rats, hypothyroidism was induced in animals by administration of methimazole (0.05%) via drinking water. After 7 weeks of treatment, hypothyroidism was confirmed by determining serum free T(3) and free T(4) concentrations. Northern blotting was used to examine the expression of PEPT2 mRNA in kidney tissues from hypothyroid rats compared to control rats. Hypothyroidism resulted in an increased level of total renal PEPT2 mRNA (121.1+/-3.3% vs. control 100+/-2.8%; p=0.008). The mRNA results were confirmed by immuno-blotting, which demonstrated significantly increased protein levels (162% vs. control 100%; p<0.01). Immunohistochemistry also revealed increased PEPT2 protein levels in the proximal tubules of treated compared to non-treated rats. In summary, PEPT2 is the first proximal tubule transporter protein that shows increased expression in states of hypothyreosis. As PEPT2 reabsorbs filtered di- and tripeptides and peptide-like drugs, the present findings may have important implications in nutritional amino acid homeostasis and for drug dynamics in states of altered thyroid function. 相似文献
10.
Summary. Glutathione (reduced form GSH and oxidized form GSSG) constitutes an important defense against oxidative stress in the brain,
and taurine is an inhibitory neuromodulator particularly in the developing brain. The effects of GSH and GSSG and glycylglycine,
γ-glutamylcysteine, cysteinylglycine, glycine and cysteine on the release of [3H]taurine evoked by K+-depolarization or the ionotropic glutamate receptor agonists glutamate, kainate, 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate
(AMPA) and N-methyl-D-aspartate (NMDA) were now studied in slices from the hippocampi from 7-day-old mouse pups in a perfusion system.
All stimulatory agents (50 mM K+, 1 mM glutamate, 0.1 mM kainate, 0.1 mM AMPA and 0.1 mM NMDA) evoked taurine release in a receptor-mediated manner. Both
GSH and GSSG significantly inhibited the release evoked by 50 mM K+. The release induced by AMPA and glutamate was also inhibited, while the kainate-evoked release was significantly activated
by both GSH and GSSG. The NMDA-evoked release proved the most sensitive to modulation: L-Cysteine and glycine enhanced the
release in a concentration-dependent manner, whereas GSH and GSSG were inhibitory at low (0.1 mM) but not at higher (1 or
10 mM) concentrations. The release evoked by 0.1 mM AMPA was inhibited by γ-glutamylcysteine and cysteinylglycine, whereas
glycylglycine had no effect. The 0.1 mM NMDA-evoked release was inhibited by glycylglycine and γ-glutamylcysteine. In turn,
cysteinylglycine inhibited the NMDA-evoked release at 0.1 mM, but was inactive at 1 mM. Glutathione exhibited both enhancing
and attenuating effects on taurine release, depending on the glutathione concentration and on the agonist used. Both glutathione
and taurine act as endogenous neuroprotective effectors during early postnatal life.
Authors’ address: Prof. Simo S. Oja, Brain Research Center, Medical School, FI-33014 University of Tampere, Finland 相似文献
11.
Summary. In the kidney the proximal tubule is responsible for the uptake of amino acids. This occurs via a variety of functionally
and structurally different amino acid transporters located in the luminal and basolateral membrane. Some of these transporters
show an ion-dependence (e.g. Na+, Cl− and K+) or use an H+-gradient to drive transport. Only a few amino acid transporters have been cloned or functionally characterized in detail
so far and their structure is known, while little is known about a majority of amino acid transporters. Only few attempts
have been untertaken looking at the regulation of amino acid transport. We summarized more recent information on amino acid
transport in the renal proximal tubule emphasizing functional and regulatory aspects.
Received August 8, 1999; Accepted April 20, 2000 相似文献
12.
Summary. 6-N-carboxymethyllysine (CML), generated by the glycation and/or oxidation of lysine residues, has been measured in biological
materials and food products using techniques such as ELISA, HPLC with fluorescence detection and mass spectrometry methods.
Only limited information has been reported regarding the preparation of standards labeled with either deuterium, 13C or 15N atoms to be used as internal standards. In the present paper, a synthesis of carbon-13 labeled CML is described using l,2-13C2-glyoxylic acid and 2-N-acetyllysine as starting materials. The resulting labeled 2-N-acetyl-CML was purified by HPLC-UV as a dibutyl ester. After a deprotection step, the yield was evaluated to be 53% when
the reaction was conducted 17 h at 37°C. CML was extensively studied by 1H- and 13C-NMR and the fragments observed in the collision induced dissociation (CID) spectrum were also assigned. Finally, the standards
of CML and carbon-13 labeled CML were accurately quantified based on 1H-NMR and tandem MS using lysine as an internal reference. 相似文献
13.
The purpose of this study was to construct stably transfected HeLa cells with human peptide transporters (hPEPT1/hPEPT2) and to identify the function of the transfected cells using the substrate JBP485 (a dipeptide) and a typical substrate for PEPTs, glycylsarcosine (Gly-Sar). An efficient and rapid method was established for the preparation and transformation of competent cells of Escherichia coli. After extraction and purification, hPEPT1/hPEPT2-pcDNA3 was transfected into HeLa cells by the liposome transfection method, respectively. HeLa-hPEPT1/hPEPT2 cells were selected by measuring the protein expression and the uptake activities of JBP485 and Gly-Sar. A simple and rapid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the simultaneous determination of JBP485 and Gly-Sar in biological samples. The Michaelis-Menten constant (K(m)) values of Gly-Sar uptake by the hPEPT1 and hPEPT2-expressing transfectants were 1.03 mM and 0.0965 mM, respectively, and the K(m) values of JBP485 uptake were 1.33 mM for PEPT1 and 0.144 mM for PEPT2. The uptake of Gly-Sar was significantly inhibited by JBP485 with a K(i) value of 8.11 mM (for PEPT1) and 1.05 mM (for PEPT2). Maximal uptake of Gly-Sar were detected at pH 5.8 (for PEPT1) and pH 6.5 (for PEPT2), suggesting that both HeLa-hPEPT1 and HeLa-hPEPT2 were H(+) dependent transporters. Stably transfected HeLa-hPEPT1/HeLa-hPEPT2 cells were constructed successfully, and the functions of hPEPT1/hPEPT2 were identified using their substrates, JBP485 and Gly-Sar. The transfected cells with transporters were used to investigate drug-drug interactions (DDIs) between JBP485 and other substrates (cephalexin or lisinopril) of PEPT1 and PEPT2. 相似文献
14.
Interaction between the actions of taurine and angiotensin II 总被引:1,自引:0,他引:1
Summary. The amino acid, taurine, is an important nutrient found in very high concentration in excitable tissue. Cellular depletion
of taurine has been linked to developmental defects, retinal damage, immundeficiency, impaired cellular growth and the development
of a cardiomyopathy. These findings have encouraged the use of taurine in infant formula, nutritional supplements and energy
promoting drinks. Nonetheless, the use of taurine as a drug to treat specific diseases has been limited. One disease that
responds favorably to taurine therapy is congestive heart failure. In this review, we discuss three mechanisms that might
underlie the beneficial effect of taurine in heart failure. First, taurine promotes natriuresis and diuresis, presumably through
its osmoregulatory activity in the kidney, its modulation of atrial natriuretic factor secretion and its putative regulation
of vasopressin release. However, it remains to be determined whether taurine treatment promotes salt and water excretion in
humans with heart failure. Second, taurine mediates a modest positive inotropic effect by regulating [Na+]i and Na+/Ca2+ exchanger flux. Although this effect of taurine has not been examined in human tissue, it is significant that it bypasses
the major calcium transport defects found in the failing human heart. Third, taurine attenuates the actions of angiotensin
II on Ca2+ transport, protein synthesis and angiotensin II signaling. Through this mechanism taurine would be expected to minimize many
of the adverse actions of angiotensin II, including the induction of cardiac hypertrophy, volume overload and myocardial remodeling.
Since the ACE inhibitors are the mainstay in the treatment of congestive heart failure, this action of taurine is probably
very important.
Received November 10, 1998, Accepted May 19, 1999 相似文献
15.
Summary. Nitric oxide (NO) has been shown to regulate neurotransmitter release in the brain; both inhibitory and excitatory effects
have been seen. Taurine is essential for the development and survival of neural cells and protects them under cell-damaging
conditions. In the brain stem, it regulates many vital functions such as cardiovascular control and arterial blood pressure.
Now we studied the effects of the NO-generating compounds hydroxylamine (HA), S-nitroso-N-acetylpenicillamine (SNAP) and sodium
nitroprusside (SNP) on the release of preloaded [3H]taurine under normal and ischemic conditions in slices prepared from the mouse brain stem from developing (7-day-old) to
young adult (3-month-old) mice. In general, the effects of NO on the release were somewhat complex and difficult to explain,
as expected from the multifunctional role of NO in the central nervous system. The basal initial release under normal conditions
was enhanced by the NO donors 5 mM HA and 1.0 mM SNAP at both ages, but SNP was inhibitory in developing mice. The release
was markedly enhanced by K+ stimulation. The effects of HA, SNAP and SNP on the basal release were not antagonized by the NO synthase inhibitor NG-nitro-L-arginine (L-NNA, 1.0 mM), demonstrating that mechanisms other than NO synthesis are involved. Taurine release in
developing mice in the presence of SNP was reduced by the inhibitor of soluble guanylate cyclase, 1H-(1,2,3)oxadiazolo(4,3-a)quinoxalin-1-one
(ODQ), indicating the possible involvement of cGMP. In normoxia, N-methyl-D-aspartate (NMDA, 1.0 mM) enhanced the SNAP- and
HA-evoked taurine release in developing mice and the HA-evoked release in adults. In ischemia, both K+ stimulation and NMDA potentiated the NO-induced release, particularly in the immature mice, probably without the involvement
of the NO synthase or cGMP. The substantial release of taurine in the developing brain stem evoked by NO donors together with
NMDA might represent signs of important mechanisms against excitotoxicity which protect the brain stem under cell-damaging
conditions.
Authors’ address: Prof. Pirjo Saransaari, Brain Research Center, Medical School University of Tampere, Tampere, FIN-3 3014,
Finland 相似文献
16.
Summary. Taurine has been thought to be essential for the development and survival of neural cells and to protect them under cell-damaging
conditions. In the brain stem taurine regulates many vital functions, including cardiovascular control and arterial blood
pressure. We have recently characterized the release of taurine in the adult and developing brain stem under normal conditions.
Now we studied the properties of preloaded [3H]taurine release under various cell-damaging conditions (hypoxia, hypoglycemia, ischemia, the presence of metabolic poisons
and free radicals) in slices prepared from the mouse brain stem from developing (7-day-old) and young adult (3-month-old)
mice, using a superfusion system. Taurine release was greatly enhanced under these cell-damaging conditions, the only exception
being the presence of free radicals in both age groups. The ischemia-induced release was characterized to consist of both
Ca2+-dependent and -independent components. Moreover, the release was mediated by Na+-, Cl−-dependent transporters operating outwards, particularly in the immature brain stem. Cl− channel antagonists reduced the release at both ages, indicating that a part of the release occurs through ion channels,
and protein kinase C appeared to be involved. The release was also modulated by cyclic GMP second messenger systems, since
inhibitors of soluble guanylyl cyclase and phosphodiesterases suppressed ischemic taurine release. The inhibition of phospholipases
also reduced taurine release at both ages. This ischemia-induced taurine release could constitute an important mechanism against
excitotoxicity, protecting the brain stem under cell-damaging conditions. 相似文献
17.
Summary. Glutaryl-CoA dehydrogenase deficiency (GDD), which is one of the most frequent organic acid disorders, is characterized by
a specific age- and regional-dependent neuropathology. We hypothesized that the distinct brain damage in GDD could be caused
by the main pathologic metabolites, the organic acids glutaric (GA) and 3-hydroxyglutaric (3-OH-GA) acids, through an excitotoxic
sequence. Therefore, we investigated the effects of 3-OH-GA and GA on primary neuronal cultures from chick embryonic telencephalons.
Here we report that 3-OH-GA and GA decreased cell viability concentration- and time-dependently, which could be only totally
prevented by preincubation with MK-801, ifenprodil and NR2B antibodies. Furthermore, cell viability decreased in parallel
with the increasing expression of NR2B subunit on cultured neurons from 2nd to 6th DIV. We conclude that GA and 3-OH-GA act as excitotoxic organic acids (EOA) specifically through NR1/NR2B and that the extent
of induced neurotoxicity is dependent on NR1/NR2B expression during maturation.
Received February 5, 1999, Accepted May 1, 1999 相似文献
18.
Taurine and neural cell damage 总被引:22,自引:2,他引:20
Summary. The inhibitory amino acid taurine is an osmoregulator and neuromodulator, also exerting neuroprotective actions in neural
tissue. We review now the involvement of taurine in neuron-damaging conditions, including hypoxia, hypoglycemia, ischemia,
oxidative stress, and the presence of free radicals, metabolic poisons and an excess of ammonia. The brain concentration of
taurine is increased in several models of ischemic injury in vivo. Cell-damaging conditions which perturb the oxidative metabolism
needed for active transport across cell membranes generally reduce taurine uptake in vitro, immature brain tissue being more
tolerant to the lack of oxygen. In ischemia nonsaturable diffusion increases considerably. Both basal and K+-stimulated release of taurine in the hippocampus in vitro is markedly enhanced under cell-damaging conditions, ischemia,
free radicals and metabolic poisons being the most potent. Hypoxia, hypoglycemia, ischemia, free radicals and oxidative stress
also increase the initial basal release of taurine in cerebellar granule neurons, while the release is only moderately enhanced
in hypoxia and ischemia in cerebral cortical astrocytes. The taurine release induced by ischemia is for the most part Ca2+-independent, a Ca2+-dependent mechanism being discernible only in hippocampal slices from developing mice. Moreover, a considerable portion of
hippocampal taurine release in ischemia is mediated by the reversal of Na+-dependent transporters. The enhanced release in adults may comprise a swelling-induced component through Cl− channels, which is not discernible in developing mice. Excitotoxic concentrations of glutamate also potentiate taurine release
in mouse hippocampal slices. The ability of ionotropic glutamate receptor agonists to evoke taurine release varies under different
cell-damaging conditions, the N-methyl-D-aspartate-evoked release being clearly receptor-mediated in ischemia. Neurotoxic
ammonia has been shown to provoke taurine release from different brain preparations, indicating that the ammonia-induced release
may modify neuronal excitability in hyperammonic conditions. Taurine released simultaneously with an excess of excitatory
amino acids in the hippocampus under ischemic and other neuron-damaging conditions may constitute an important protective
mechanism against excitotoxicity, counteracting the harmful effects which lead to neuronal death. The release of taurine may
prevent excitation from reaching neurotoxic levels.
Received January 25, 2000/Accepted January 31, 2000 相似文献
19.
Maize (Zea mays L.) cell cultures incorporated radioactivity from [14C]cinnamate into hydroxycinnamoyl-CoA derivatives and then into polysaccharide-bound feruloyl residues. Within 5–20 min, the
CoA pool had lost its 14C by turnover and little or no further incorporation into polysaccharides then occurred. The system was thus effectively a
pulse–chase experiment. Kinetics of radiolabelling of diferulates (also known as dehydrodiferulates) varied with culture age.
In young (1–3 d) cultures, polysaccharide-bound [14C]feruloyl- and [14C]diferuloyl residues were both detectable within 1 min of [14C]cinnamate feeding. Thus, feruloyl residues were dimerised <1 min after their attachment to polysaccharides. For at least
the first 2.3 h after [14C]cinnamate feeding, polysaccharide-bound [14C]diferuloyl residues remained almost constant at ≈7% of the total polysaccharide-bound [14C]ferulate derivatives. Since feruloyl residues are attached to polysaccharides <1 min after the biosynthesis of the latter,
and >10 min before secretion, the data show that extensive feruloyl coupling occurred intra-protoplasmically. Exogenous H2O2 (1 mM) caused little additional feruloyl coupling; therefore, wall-localised coupling may have been peroxidase-limited. In
older (e.g. 4 d) cultures, less intraprotoplasmic coupling occurred: during the first 2.5 h, polysaccharide-bound [14C]diferuloyl residues were a steady 1.4% of the total polysaccharide-bound [14C]ferulate derivatives. In contrast to the situation in younger cultures, exogenous H2O2 induced a rapid 4- to 6-fold increase in all coupling products, indicating that coupling in the walls was H2O2-limited. In both 2- and 4-d-old cultures, polysaccharide-bound 14C-trimers and larger coupling products exceeded [14C]diferulates 3- to 4-fold, but followed similar kinetics. Thus, although all known dimers of ferulate can now be individually
quantified, it appears to be trimers and larger products that make the major contribution to cross-linking of wall polysaccharides
in cultured maize cells. We argue that feruloyl arabinoxylans that are cross-linked before and after secretion are likely
to loosen and tighten the cell wall, respectively. The consequences for the control of cell expansion and for the response
of cell walls to an oxidative burst are discussed.
Received: 19 January 2000 / Accepted: 13 April 2000 相似文献
20.
Electrophysiological characteristics of the proton-coupled peptide transporter PEPT2 cloned from rat brain 总被引:7,自引:0,他引:7
Wang Hong; Fei You-Jun; Ganapathy Vadivel; Leibach Frederick H. 《American journal of physiology. Cell physiology》1998,275(4):C967
We have cloned a peptide transporter from rat brain and found itto be identical to rat kidney PEPT2. In the present study wecharacterize the transport function of the rat brain PEPT2, withspecial emphasis on electrophysiological properties and interaction withN-acetyl-L-aspartyl-L-glutamate(NAAG). When heterologously expressed in HeLa cells and in SK-N-SHcells, PEPT2 transports several dipeptides but not free amino acids inthe presence of a proton gradient. NAAG competes with other peptidesfor the PEPT2-mediated transport process. When PEPT2 is expressed inXenopus laevis oocytes, substrate-induced inward currents are detectable with dipeptides ofdiffering charge in the presence of a proton gradient. Proton activation kinetics are similar for differently charged peptides. NAAGis a transportable substrate for PEPT2, as evidenced by NAAG-induced currents. The Hill coefficient for protons for the activation of thetransport of differently charged peptides, including NAAG, is 1. Although the peptide-to-proton stoichiometry for negatively chargedpeptides is 1, the transport nonetheless is associated with transfer ofpositive charge into the oocyte, as indicated by peptide-induced inwardcurrents. 相似文献