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1.
A genetic locus controlling the electrophoretic mobility of an acid phosphatase in mouse kidney is described. This locus, called acid phosphatase-kidney (Apk), is not expressed in erythrocytes, liver, spleen, heart, lung, brain, skeletal muscle, stomach, or testes. The product of Apk hydrolyzes the substrate naphthol AS-MX phosphoric acid but is not active on a-naphthylphosphate or 4-methylumbelliferylphosphate. It is not inactivated by 50 C for 1 hr, nor is its electrophoretic mobility altered by incubation with neuraminidase. The locus is invariant among 31 inbred strains (Apk
a), with a variant allele (Apk
m) observed only in Mus musculus molossinus. Codominant expression was observed in F1 hybrids of M. m. molossinus and inbred strains. Apk was mapped on Chr 10, near the neurological mutant waltzer (v).This work was supported by Contract NO1-ES42159 from the National Institute of Environmental Health Sciences and by Grant 1-476 from the National Foundation—March of Dimes. The Jackson Laboratory is fully accredited by the American Association for Accreditation of Laboratory Animal Care. 相似文献
2.
Jeffrey C. Gingrich Jeffrey S. Buzby Veronica L. Stirewalt Donald A. Bryant 《Photosynthesis research》1988,16(1-2):83-99
Two herbicide-resistant strains of the cyanobacterium Synechococcus sp. PCC 7002 are compared to the wild-type with respect to the DNA changes which result in herbicide resstance. The mutations have previously been mapped to a region of the cyanobacterial genome which encodes oneof three copies of psbA, the gene which encodes the 32 kDa Qb-binding protein also known as D1 (Buzby et al. 1987). The DNA sequence of the wild-type gene was first determined and used as a comparison to that of the mutant alleles. A point mutation at codon 211 in the psbA1 coding locus (TTC) to TCC) results in an amino acid change from phenylalanine to serine in the D1 protein. This mutation confers resistance to atrazine and diuron at seven times and at two times the minimal inhibitory concentration (MIC) for the wild-type, respectively. A mutation at codon 211 resulting in herbicide resistance has not previously been described in the literature. A second point mutation at codon 219 in the psbA1 coding locus (GTA to ATA) results in an amino acid change from valine to isoleucine in the D1 protein. This mutation confers resistance to diuron and atrazine at ten times and at two times the MIC for the wild-type, respectively. An identical codon change conferring similar herbicide resistance patterns has previously been described in Chlamydomonas reinhardtii. The atrazine-resistance phenotype in Synechococcus sp. PCC 7002 was shown to be dominant by plasmid segregation analysis.Abbreviations At
r
atrazine resistance
- Du
r
diuron resistance
- Km
r
kanamycin resistance
- Ap
r
ampicillin resistance
- MIC
Minimum inhibitory concentration 相似文献
3.
Jee Won Im Nam Deuk Kim Nam Deuk Kim Byung Pal Yu Hee-Sun Yang Hae Young Chung 《Biotechnology letters》2005,26(21):1665-1669
The arachidonate cascade is important for the generation of reactive species (RS), and cyclooxygenase (COX) is a key enzyme of this cascade. Tissues of 24-month-old rat lung showed a 2-fold increase in RS, malondialdehyde and thromboxane B2 than those of 6-month-old rat. We found that the effects of 50 µM H2O2 and 200 µM
t-butylhydroperoxide (t-BHP) specify on COX activity, and that their effects increased cytosolic COX activity in a concentration-dependent manner (1–50 µM) in 24-month-old rat. Our results suggested that COX activators such as t-BHP and H2O2, which are located in cytosol, are essential for the activation of COX in aged lung. 相似文献
4.
Yasuhiro Hashimoto Mikiko Abe Akemi Suzuki Kentaro Iwasaki Tamio Yamakawa 《Glycoconjugate journal》1985,2(3-4):255-265
Genetic polymorphism in the expression of the GM1(NeuGc) ganglioside has been shown in the liver of inbred strains of mice. Through analysis of the gangliosides of H-2 congenic and recombinant strains, this polymorphism was demonstrated to be controlled by a locus mapped left outside of the H-2 complex on chromosome 17, and the locus was assumed to control the level of the activity of GM1(NeuGc) synthetase, UDP-galactose:GM2(NeuGc) galactosyltransferase (E.C.2.4.1.62) [Hashimotoet al., J Biochem (1983) 94:2049-54].In the present study we analyzed the genetic linkage between the activity of the galactosyltransferase and the H-2 haplotype. For this purpose, we selected two inbred strains of mice, WHT/Ht and BALB/c, because they have different levels of the transferase activity and show different H-2 haplotypes; the specific activity of the transferase obtained with BALB/c was one-eighth of that with WHT/Ht, and BALB/c expressed the la.7 antigen as one of the products encoded in their H-2d complex, whereas WHT/Ht did not. To analyze the linkage between these two phenotypes, WHT/Ht were mated with BALB/c to obtain the F1 mice, and the female F1 mice were then backcrossed to WHT/Ht. It was found that one half of the backcross generation expressed the la.7 antigen derived from BALB/c and had a significantly lower specific activity of the transferase than that of WHT/Ht, while the other half did not express the la.7 antigen but had the same specific activity of the transferase as that obtained with WHT/Ht.These results suggest that the locus controlling the level of the transferase activity in mouse liver is linked to the H-2 complex on chromosome 17.Abbreviations NeuGc
N-glycolylneuraminic acid
The ganglioside nomenclature is based on the system of Svennerholm, J Neurochem (1963) 10:613-23. The sialic acid species present is shown in parentheses after the ganglioside abbreviation. 相似文献
5.
Dorothee Heuermann Peter Roggentin Reinhard G. Kleineidam Roland Schauer 《Glycoconjugate journal》1991,8(2):95-101
The sialidase secreted byClostridium chauvoei NC08596 was purified to apparent homogeneity by ion-exchange chromatography, gel filtration, hydrophobic interaction-chromatography, FPLC ion-exchange chromatography, and FPLC gel filtration. The enzyme was enriched about 10 200-fold, reaching a final specific activity of 24.4 U mg–1. It has a relatively high molecular mass of 300 kDa and consists of two subunits each of 150 kDa. The cations Mn2+, Mg2+, and Ca2+ and bovine serum albumin have a positive effect on the sialidase activity, while Hg2+, Cu2+, and Zn2+, chelating agents and salt decrease enzyme activity. The substrate specificity, kinetic data, and pH optimum of the enzyme are similar to those of other bacterial sialidases.Abbreviations FPLC
fast protein liquid chromatography
- NCTC
National Collection of Type Cultures
- ATCC
American Type Culture Collection
- MU-Neu5Ac
4-methylumbelliferyl--d-N-acetylneuraminic acid
- buffer A
0.02m piperazine, 0.01m CaCl2, pH 5.5
- buffer B
0.02m piperazine, 0.01m CaCl2, 1.0m NaCl, pH 5.5
- buffer C
0.1m sodium acetate, 0.01m CaCl2, pH 5.5
- SDS
sodium dodecyl sulfate
- PAGE
polyacrylamide gel electrophoresis
- Neu5Ac
N-acetylneuraminic acid
- BSM
bovine submandibular gland mucin
- GD1a
IV3Neu5Ac, II3Neu5Ac-GgOse4Cer
- GM1
II3Neu5Ac-GgOse4Cer
- MU-Neu4,5Ac2
4-methylumbelliferyl--d-N-acetyl-4-O-acetylneuraminic acid
- TLC
thin-layer chromatography
- HPTLC
high performance thin-layer chromatography
- EDTA
ethylenediamine tetraacetic acid
- EGTA
ethylene glycol bis(2-aminoethyl-ethen)-N,N,N,N-tetraacetic acid
- BSA
bovine serum albumin
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid
- IEF
isoelectric focusing
- IEP
isoelectric point 相似文献
6.
Birgitta M. W?hrl Udo F. Wehmeier Joseph W. Lengeler 《Molecular & general genetics : MGG》1990,224(2):193-200
Summary In Klebsiella pneumoniae the gene products involved in the degradation of the ketose l-sorbose are encoded in the sor operon. It comprises, besides structural genes for uptake and catabolism, a promoter-proximal gene sorC, encoding a protein SorC of Mr 40 kDa, for which no enzymatic function has been detected. All sor genes are coordinately expressed and inducible by l-sorbose. Polar insertions and frameshift mutations in sorC cause a pleiotropic negative effect on the expression of all other sor genes. This defect is complemented in trans by the wild-type sorC
+ allele for frameshift mutations, but not for polar insertions. A single promoter for all sor genes, for which SorC is the activator, thus seems to be located in front of sorC. The repressor activity of SorC was demonstrated by complementation of constitutive sorC alleles with a sorC
+ allele leading to inducible expression of all sor genes, including sorC, which, as visualized by the use of a series of lacZ fusions, thus autoregulates its expression, both as an activator and a repressor. 相似文献
7.
Gabriele D'andrea Jan B Bouwstra Johannis P Kamerling Johannes F G Vliegenthart 《Glycoconjugate journal》1988,5(2):151-157
Ascorbic acid oxidase (E.C.1.10.3.3) from the green zucchini squash (Cucurbita pepo medullosa) is a copper-containing glycoprotein which catalyzes the reaction:l-ascorbic acid +1/2 O2l-dehydroascorbic acid + H2O. The carbohydrate content of the purified plant glycoprotein amounted to 3% (w/w), and monosaccharide analysis revealed the carbohydrate moiety to be of theN-glycosidic type. The carbohydrate chains were released from the apoenzyme by digestion with PNGase-F immobilized on Sepharose 4B. After fractionation on Bio-Gel P-2 and purification on Mono-Q, the neutral oligosaccharide was investigated by 500-MHz1H-NMR spectroscopy. The primary structure of theN-linked carbohydrate chain was established to be:
Abbreviations AAO
ascorbic acid oxidase
- PNGase-F
peptide-N
4-(N-acetyl--glucosaminyl)asparagine amidase-F
- GalNAc
N-acetylgalactosamine
- GlcNAc
N-acetylglucosamine
- Man
mannose
- Xyl
xylose
- GLC
gas-liquid chromatography
- FPLC
fast protein liquid chromatography
- NMR
nuclear magnetic resonance
- SDS-PAGE
sodium dodecyl sulfate-polyacrylamide gel electrophoresis 相似文献
8.
Evelyn A. Havir 《Planta》1981,152(2):124-130
Suspension-cultured cells of soybean (Glycine max (L.) Merr. cv. Kanrich) produce large amounts of phenylalanine ammonia-lyase (PAL; EC 4.3.1.5), the first enzyme of phenylpropanoid metabolism, during growth. 2-Aminooxyacetic acid (AOA) and l-2-aminooxy-3-phenylpropionic acid (l-AOPP) inhibit the enzyme competitively in vitro and have been used for in vivo studies. The amount of extractable enzyme in the cells and their utilization of NO
3
–
and NH
3
+
are reduced upon the addition of AOA. When AOA was added at various times during growth, the appearance of additional enzyme activity was prevented but enzyme already formed was not inhibited. No evidence was obtained for the presence of an inhibitor in the extracts and AOA inhibition in vitro was readily reversible. It is conculded that AOA acts to inhibit the formation of PAL in suspension-cultured soy bean cells. In vitro inhibition of soybean PAL by l-AOPP could not be reversed; in contrast, the inhibition of maize (Zea mays L.) PAL was readily reversible. Added l-AOPP, which was rapidly taken up by the soybean cells, prevented the large increase in enzyme activity. Although PAL activity was blocked in the cultures, no appreciable increase in phenylalanine content could be detected in cell extracts. The response of soybean cell suspensions to l-AOPP addition thus differs from that of other tissues which in presence of l-AOPP show an increase in PAL activity and an accumulation of phenylalanine.Abbreviations AOA
2-aminooxyacetic acid
-
l-AOPP
l-2-aminoxy-3-phenylpropionic acid
- PAL
l-phenylalanine ammonialyase (EC4.3.1.5) 相似文献
9.
VGLUT2 is one of three vesicular glutamate transporters that play crucial roles in glutamatergic excitatory neurotransmission.
We explored the functional properties of the rat VGLUT2 by heterologous expression of VGLUT2 in Xenopus oocytes. Immunocytochemical analysis indicated that most VGLUT2 protein was expressed in intracellular compartments but that
some expression occurred also on the plasma membrane. Functional analysis revealed VGLUT2 to be active in two independent
modes, namely, uptake into intracellular organelles and efflux at the plasma membrane. VGLUT-specific transport was identified
based on the strong preference for glutamate over aspartate—in contrast to plasma-membrane or mitochondrial glutamate transporters—and
sensitivity to known VGLUT blockers. VGLUT2 expression in oocytes (1) stimulated the influx of l-[3H]glutamate, but not d-[3H]aspartate, into digitonin-permeabilized oocytes and (2) stimulated efflux of l-glutamate, but not l-aspartate, from intact oocytes preinjected with 3H-labeled amino acids. In the latter assay, cellular efflux of glutamate (which was blocked by rose bengal and trypan blue)
may be analogous to vesicular packaging of glutamate. Our data are consistent with VGLUT2-mediated H+/l-glutamate antiport, but not antiport with chloride. Expression of mammalian VGLUT1 and VGLUT3 also stimulated l-[3H]glutamate efflux from Xenopus oocytes, suggesting that this phenomenon is a general feature of vesicular glutamate transporters. Our findings support the
idea that vesicular glutamate transporters, when transiently expressed on the neuronal plasma membrane, may mediate Ca2+-independent glutamate leakage in addition to their traditional role of packaging glutamate into synaptic vesicles for Ca2+-dependent exocytosis.
Special issue article in honor of Dr. Frode Fonnum. 相似文献
10.
On following N2-incorporation and subsequent metabolism in the lichen Peltigera canina using 15N as tracer, it was found, over a 30 min period, that greatest initial labelling was into NH
4
+
followed by glutamate and the amide-N of glutamine. Labelling of the amino-N of glutamine, aspartate and alanine increased slowly. Pulse-chase experiments using 15N confirmed this pattern. On inhibiting the GS-GOGAT pathway using l-methionine-dl-sulphoximine and azaserine, 15N enrichment of glutamate, alanine and aspartate continued although labelling of glutamine was undetectable. From this and enzymic data, NH
4
+
assimilation in the P. canina thallus appears to proceed via GS-GOGAT in the cyanobacterium and via GDH in the fungus; aminotransferases were present in both partners. The cyanobacterium assimilated 44% of the 15N2 fixed; the remainder was liberated almost exclusively as NH
4
+
and then assimilated by fungal GDH.Abbreviations ADH
alanine dehydrogenase
- APT
aspartate-pyruvate aminotransferase
- AOA
aminooxyacetate
- GDH
glutamate dehydrogenase
- GOT
glutamate-oxaloacetate aminotransferase
- GOGAT
glutamate synthase
- GPT
glutamate-pyruvate aminotransferase
- GS
glutamine synthetase
- HEPES
4-(2-hydroxyethyl)-1-piperazine ethanesulphonic acid
- MSX
l-methionine-dl-sulphoximine 相似文献
11.
Ugarte G Delgado R O'Day PM Farjah F Cid LP Vergara C Bacigalupo J 《The Journal of membrane biology》2005,207(3):151-160
We report that Drosophila retinal photoreceptors express inwardly rectifying chloride channels that seem to be orthologous to mammalian ClC-2 inward
rectifier channels. We measured inwardly rectifying Cl− currents in photoreceptor plasma membranes: Hyperpolarization under whole-cell tight-seal voltage clamp induced inward Cl− currents; and hyperpolarization of voltage-clamped inside-out patches excised from plasma membrane induced Cl− currents that have a unitary channel conductance of ∼3.7 pS. The channel was inhibited by 1 mM Zn2+ and by 1 mM 9-anthracene, but was insensitive to DIDS. Its anion permeability sequence is Cl− = SCN−> Br−>> I−, characteristic of ClC-2 channels. Exogenous polyunsaturated fatty acid, linolenic acid, enhanced or activated the inward
rectifier Cl− currents in both whole-cell and excised patch-clamp recordings. Using RT-PCR, we found expression in Drosophila retina of a ClC-2 gene orthologous to mammalian ClC-2 channels. Antibodies to rat ClC-2 channels labeled Drosophila photoreceptor plasma membranes and synaptic regions. Our results provide evidence that the inward rectification in Drosophila retinal photoreceptors is mediated by ClC-2-like channels in the non-transducing (extra-rhabdomeral) plasma membrane, and
that this inward rectification can be modulated by polyunsaturated fatty acid.
G. Ugarte and R. Delgado contributed equally to this work. 相似文献
12.
Paweł Wiśniewski Justyna Szklarczyk Marcin Maciaga Andrzej Paszkowski 《Acta Physiologiae Plantarum》2006,28(6):577-588
The occurrence of four l-alanine:2-oxoglutarate aminotransferase (AOAT) isoenzymes (AOAT-like proteins): alanine aminotransferase 1 and 2 (AlaAT1
and AlaAT2, EC 2.6.1.2) and l-glutamate:glyoxylate aminotransferase 1 and 2 (GGAT1 and GGAT2, EC 2.6.1.4) was demonstrated in Arabidopsis thaliana leaves. These enzymes differed in their substrate specificity, susceptibility to pyridoxal phosphate inhibitors and behaviour
during molecular sieving on Zorbax SE-250 column. A difference was observed in the electrostatic charge values at pH 9.1 between
GGAT1 and GGAT2 as well as between AlaAT1 and AlaAT2, despite high levels of amino acid sequence identity (93 % and 85 %,
respectively). The unprecedented evidence for the monomeric structure of both AlaAT1 and AlaAT2 is presented. The molecular
mass of each enzyme estimated by molecular sieving on Sephadex G-150 and Zorbax SE-250 columns and SDS/PAGE was approximately
60 kDa. The kinetic parameters: Km (Ala)=1.53 mM, Km (2-oxoglutarate)=0.18 mM, kcat=124.6 s−1, kcat/Km=8.1 × 104 M−1·s−1 of AlaAT1 were comparable to those determined for other AlaATs isolated from different sources. The two studied GGATs also
consisted of a single subunit with molecular mass of 47.3–70 kDa. The estimated Km values for l-glutamate (1.2 mM) and glyoxylate (0.42 mM) in the transamination catalyzed by putative GGAT1 contributed to indentification
of the enzyme. Based on these results we concluded that each of four AOAT genes in Arabidopsis thaliana leaves expresses different AOAT isoenzyme, functioning in a native state as a monomer. 相似文献
13.
An epoxide hydrolase gene of about 0.8 kb was cloned from Rhodococcus opacus ML-0004, and the open reading frame (ORF) sequence predicted a protein of 253 amino acids with a molecular mass of about
28 kDa. An expression plasmid carrying the gene under the control of the tac promotor was introduced into Escherichia coli, and the epoxide hydrolase gene was successfully expressed in the recombinant strains. Some characteristics of purified recombinant
epoxide hydrolase were also studied. Epoxide hydrolase showed a high stereospecificity for l(+)-tartaric acid, but not for d(+)-tartaric acid. The epoxide hydrolase activity could be assayed at the pH ranging from 3.5 to 10.0, and its maximum activity
was obtained between pH 7.0 and 7.5. The enzyme was sensitive to heat, decreasing slowly between 30°C and 40°C, and significantly
at 45°C. The enzyme activity was activated by Ca2+ and Fe2+, while strongly inhibited by Ag+ and Hg+, and slightly inhibited by Cu2+, Zn2+, Ba2+, Ni+, EDTA–Na2 and fumarate. 相似文献
14.
1-Aminocyclopropane-1-carboxylic acid (ACC) is known to be converted to ethylene and conjugated into N-malonyl-ACC in plant tissues. When -amino[1-14C]isobutyric acid (AIB), a structural analog of ACC, was administered to mungbean (Vigna radiata L.) hypocotyl segments, it was metabolized to 14CO2 and conjugated to N-malonyl-AIB (MAIB). -Aminoisobutyric acid inhibited the conversion of ACC to ethylene and also inhibited, to a lesser extent, N-malonylation of ACC and d-amino acids. Although the malonylation of AIB was strongly inhibited by ACC as well as by d-amino acids, the metabolism of AIB to CO2 was inhibited only by ACC but not by d-amino acids. Inhibitors of ACC conversion to ethylene such as anaerobiosis, 2,4-dinitrophenol and Co2+, similarly inhibited the conversion of AIB to CO2. These results indicate that the malonyalation of AIB to MAIB is intimately related to the malonylation of ACC and d-amino acids, whereas oxidative decarboxylation of AIB is related to the oxidative degradation of ACC to ethylene.Abbreviations ACC
1-aminocyclopropane-1-carboxylic acid
- AIB
-aminoisobutyric acid
- MACC
1-(malonylamino)-cyclopropane-1-carboxylic acid
- MAIB
-(malonylamino)-isobutyric acid
- Mes
2-(N-morpholino)ethanesulfonic acid 相似文献
15.
Functional expression of rat renal Na/Pi-cotransport (NaPi-2) in Sf9 cells by the baculovirus system
The recently cloned Na/P
i
-cotransport system NaPi-2 is an apical membrane protein of rat proximal tubular cells and is involved in proximal phosphate reabsorption. To make the protein available for further functional/structural studies, this transport system has been expressed in Sf9 insect cells using a recombinant baculovirus. Sf9 cells infected with NaPi-2 (or 6His tagged NaPi-2) expressed functional Na/P
i
-cotransport up to 20- to 50-fold over noninfected Sf9 cells. Transport of phosphate in infected cells was highly dependent on sodium, exhibited a K
m
for P
i
of 0.114 mm and an apparent K
m
for Na of 63 mm (Hill coefficient of approximately 3) and was stimulated by high external pH. Infected cells expressed a polypeptide of 65 kDa representing a nonglycosylated form of the 85 kDa mature NaPi-2 transporter as present in proximal tubular brush-border membranes. By confocal microscopy expression of NaPi-2 protein was observed in the plasma membrane, yet submembranous accumulation of NaPi-2 protein could not be excluded. This demonstrates that the rat proximal tubular Na/P
i
-cotransport system NaPi-2 can be successfully expressed in Sf9 cells with characteristics similar to that in isolated brush-border membranes. The 6His tagging will permit isolation of the NaPi-2 cotransporter in amounts sufficient for structural/functional studies.We would like to thank W. Scherle and M. Lötscher (Institute of Anatomy) for their generous help using the confocal microscope and Ch. Gasser for the art work. Financial support by the Swiss National Fonds [Grant No. 32-30785.91 (to H.M.) and 32-28664.90 (to J.B.)] and by Stiftung für wissenschaftliche Forschung an der Universitát Zürich is greatly acknowledged. 相似文献
16.
Guennoun-Lehmann S Fonseca JE Horisberger JD Rakowski RF 《The Journal of membrane biology》2007,216(2-3):107-116
Palytoxin (PTX) opens a pathway for ions to pass through Na,K-ATPase. We investigate here whether PTX also acts on nongastric
H,K-ATPases. The following combinations of cRNA were expressed in Xenopus laevis oocytes: Bufo marinus bladder H,K-ATPase α2- and Na,K-ATPase β2-subunits; Bufo Na,K-ATPase α1- and Na,K-ATPase β2-subunits; and Bufo Na,K-ATPase β2-subunit alone. The response to PTX was measured after blocking endogenous Xenopus Na,K-ATPase with 10 μm ouabain. Functional expression was confirmed by measuring 86Rb uptake. PTX (5 nm) produced a large increase of membrane conductance in oocytes expressing Bufo Na,K-ATPase, but no significant increase occurred in oocytes expressing Bufo H,K-ATPase or in those injected with Bufo β2-subunit alone. Expression of the following combinations of cDNA was investigated in HeLa cells: rat colonic H,K-ATPase α1-subunit and Na,K-ATPase β1-subunit; rat Na,K-ATPase α2-subunit and Na,K-ATPase β2-subunit; and rat Na,K-ATPase β1- or Na,K-ATPase β2-subunit alone. Measurement of increases in 86Rb uptake confirmed that both rat Na,K and H,K pumps were functional in HeLa cells expressing rat colonic HKα1/NKβ1 and NKα2/NKβ2. Whole-cell patch-clamp measurements in HeLa cells expressing rat colonic HKα1/NKβ1 exposed to 100 nm PTX showed no significant increase of membrane current, and there was no membrane conductance increase in HeLa cells transfected
with rat NKβ1- or rat NKβ2-subunit alone. However, in HeLa cells expressing rat NKα2/NKβ2, outward current was observed after pump activation by 20 mm K+ and a large membrane conductance increase occurred after 100 nm PTX. We conclude that nongastric H,K-ATPases are not sensitive to PTX when expressed in these cells, whereas PTX does act
on Na,K-ATPase. 相似文献
17.
Two types of alleles exist in the human alcohol dehydrogenase-2 (ADH
2) locus. The usualADH
2
1
allele is common in Caucasians, while the atypicalADH
2
2
allele is predominant in Orientals. TheADH
2
2
produces the β2 subunit, which is catalytically far more active than the β1 subunit produced by theADH
2
1
gene. The racial difference in alcohol-related problems could be related to the genetic differences in ADH and other ethanol-metabolizing
enzymes. In order to examine the possibility, a method for determiningADH
2 genotypes was developed. Two 21-base synthetic oligonucleotides, one complementary to the usualADH
2
1
allele and the other complementary to the atypicalADH
2
2
allele, were used as specific probes for in-gel hybridization analysis of human genomic DNA from peripheral blood. Under
appropriate hybridization conditions, these two probes can hybridize to their specific complementary alleles and, thus, allow
the genotyping of theADH
2 locus. Genotypes of theADH
2 locus of 49 unrelated Japanese individuals were determined. The frequency of the atypicalADH
2
2
gene was found to be 0.71 in the Japanese population examined.
This research was supported by Grant AA05763 from the National Institutes of Health. 相似文献
18.
Plantaricin A (PlnA) is a 26-mer peptide pheromone with membrane-permeabilizing, strain-specific antibacterial activity, produced
by Lactobacillus plantarum C11. We investigated the membrane-permeabilizing effects of PlnA on cultured cancerous and normal rat anterior pituitary
cells using patch-clamp techniques and microfluorometry (fura-2). Cancerous cells displayed massive permeabilization within
5 s after exposure to 10–100 μm PlnA. The membrane depolarized to nearly 0 mV, and the membrane resistance decreased to a mere fraction of the initial value after less than 1 min. In outside-out membrane
patches, 10 μm PlnA induced membrane currents reversing at 0 mV, which is compatible with an unspecific conductance increase. The d and l forms of the peptide had similar potency, indicating a nonchiral mechanism for the membrane-permeabilizing effect. Surprisingly,
inside-out patches were insensitive to 1 mm PlnA. Primary cultures of normal rat anterior pituitary cells were also insensitive to the peptide. Thus, PlnA differentiates
between plasma membranes and membrane leaflets. Microfluorometric recordings of [Ca2+]
i
and cytosolic concentration of fluorochrome verified the rapid permeabilizing effect of PlnA on cancerous cells and the insensitivity
of normal pituitary cells. 相似文献
19.
During L-lactic acid fermentation by Rhizopus oryzae, increasing the phosphate level in the fermentation medium from 0.1 g l–1 to 0.6 g l–1 KH2PO4 reduced the maximal concentration of L-lactic acid and fumaric acid from 85 g l–1 to 71 g l–1 and from 1.36 g l–1 to 0.18 g l–1, respectively; and it decreased the fermentation time from 72 h to 52 h. Phosphate at 0.40 g l–1 KH2PO4 was suitable for both minimizing fumaric acid accumulation and benefiting L-lactic acid production. 相似文献
20.
Stimulant-evoked depolarization and increase in [Ca2+]
i
in insulin-secreting cells is dependent on external Na+ 总被引:1,自引:0,他引:1
M. J. Dunne D. I. Yule D. V. Gallacher O. H. Petersen 《The Journal of membrane biology》1990,113(2):131-138
Summary The patch-clamp technique and measurements of single cell [Ca2+]
i
have been used to investigate the importance of extracellular Na+ for carbohydrate-induced stimulation of RINm5F insulin-secreting cells. Using patch-clamp whole-cell (current-clamp) recordings the average cellular transmembrane potential was estimated to be –60±1 mV (n=83) and the average basal [Ca2+]
i
102±6nm (n=37). When challenged with either glucose (2.5–10mm) ord-glyceraldehyde (10mm) the cells depolarized, which led to the initiation of Ca2+ spike potentials and a sharp rise in [Ca2+]
i
. Similar effects were also observed with the sulphonylurea compound tolbutamide (0.01–0.1mm). Both the generation of the spike potentials and the increase in [Ca2+]
i
were abolished when Ca2+ was removed from the bathing media. When all external Na+ was replaced with N-methyl-d-glucamine, in the continued presence of either glucose,d-glyceraldehyde or tolbutamide, a membrane repolarization resulted, which terminated Ca2+ spike potentials and attenuated the rise in [Ca2+]
i
. Tetrodotoxin (TTX) (1–2 m) was also found to both repolarize the membrane and abolish secretagogue-induced rises in [Ca2+]
i
. 相似文献