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61.
The initial rate of Fe(2+) movement across the inner envelope membrane of pea (Pisum sativum) chloroplasts was directly measured by stopped-flow spectrofluorometry using membrane vesicles loaded with the Fe(2+)-sensitive fluorophore, Phen Green SK. The rate of Fe(2+) transport was rapid, coming to equilibrium within 3s. The maximal rate and concentration dependence of Fe(2+) transport in predominantly right-side-out vesicles were nearly equivalent to those measured in largely inside-out vesicles. Fe(2+) transport was stimulated by an inwardly directed electrochemical proton gradient across right-side-out vesicles, an effect that was diminished by the addition of valinomycin in the presence of K(+). Fe(2+) transport was inhibited by Zn(2+), in a competitive manner, as well as by Cu(2+) and Mn(2+). These results indicate that inward-directed Fe(2+) transport across the chloroplast inner envelope occurs by a potential-stimulated uniport mechanism. 相似文献
62.
Presenilins play prominent roles in the molecular pathogenesis of Alzheimer's disease and during embryo development. We have isolated a zebrafish presenilin orthologue (pre2), which shows a high degree of sequence identity to the human PS2 protein. Zebrafish pre2 is maternally and ubiquitously expressed during early embryo development, whereas Pre2 protein expression is initiated between 6 and 12 hours post fertilisation (hpf), suggesting strict regulation of pre2 translation. pre2 expression is especially high in neural-crest-derived melanocytes. 相似文献
63.
64.
The epsilon subunit of the chloroplast ATP synthase is an inhibitor of activity of the enzyme. Recombinant forms of the epsilon subunit from spinach chloroplasts lacking the last 10, 32, or 45 amino acids were immobilized onto activated Sepharose. A polyclonal antiserum raised against the epsilon subunit was passed over these immobilized protein columns, and the purified antibodies which were not bound recognized the portions of the epsilon subunit missing from the recombinant form present on the column. The full polyclonal antiserum can strip the epsilon subunit from the ATP synthase in illuminated thylakoid membranes [Richter, M. L., and McCarty, R. E. (1987) J. Biol. Chem. 262, 15037-15040]. Exposure of illuminated thylakoid membranes to antibodies recognizing the last 32 amino acids of the epsilon subunit collapses the proton gradient and hinders ATP synthesis with similar efficiency as the full polyclonal preparation. These results indicate that antibodies against the last 32 amino acids of the epsilon subunit are capable of stripping the subunit from the ATP synthase in illuminated membranes. Neither of these effects was seen when the membranes were exposed to the antibodies in the dark. This is direct evidence that the chloroplast ATP synthase undergoes a conformational shift during its activation by the electrochemical proton gradient which specifically alters the conformation of the carboxyl-terminal domain of the epsilon subunit from protected to solvent-exposed. The relation between this shift and activation of the enzyme by the electrochemical proton gradient is discussed. 相似文献
65.
66.
Blockers of CFTR with well-characterized kinetics and mechanism of action will be useful as probes of pore structure. We
have studied the mechanism of block of CFTR by the arylaminobenzoates NPPB and DPC. Block of macroscopic currents by NPPB
and DPC exhibited similar voltage-dependence, suggestive of an overlapping binding region. Kinetic analysis of single-channel
currents in the presence of NPPB indicate drug-induced closed time constants averaging 2.2 msec at −100 mV. The affinity for
NPPB calculated from single-channel block, K
D
= 35 μm, exceeds that for other arylaminobenzoates studied thus far. These drugs do not affect the rate of activation of wild-type
(WT) channels expressed in oocytes, consistent with a simple mechanism of block by pore occlusion, and appear to have a single
binding site in the pore. Block by NPPB and DPC were affected by pore-domain mutations in different ways. In contrast to its
effects on block by DPC, mutation T1134F-CFTR decreased the affinity and reduced the voltage-dependence for block by NPPB.
We also show that the alteration of macroscopic block by NPPB and DPC upon changes in bath pH is due to both direct effects
(i.e., alteration of voltage-dependence) and indirect effects (alteration of cytoplasmic drug loading). These results indicate
that both NPPB and DPC block CFTR by entering the pore from the cytoplasmic side and that the structural requirements for
binding are not the same, although the binding regions within the pore are similar. The two drugs may be useful as probes
for overlapping regions in the pore.
Received: 14 October 1999/Revised: 18 January 2000 相似文献
67.
The chloroplast ATP synthase is strictly regulated so that it is very active in the light (rates of ATP synthesis can be higher than 5 mol/min/mg protein), but virtually inactive in the dark. The subunits of the catalytic portion of the ATP synthase involved in activation, as well as the effects of nucleotides are discussed. The relation of activation to proton flux through the ATP synthase and to changes in the structure of enzyme induced by the proton electrochemical gradient are also presented. It is concluded that the and subunits of CF1 play key roles in both regulation of activity and proton translocation. 相似文献
68.
Leaf senescence-like characteristics contribute to cotton's premature photosynthetic decline 总被引:6,自引:0,他引:6
Leaf and canopy photosynthesis of cotton (Gossypium hirsutum L.) declines as the crop approaches cutout, just as the assimilate needs for reproductive growth are peaking. Our objective
with this study was to determine whether this decline is due to remobilization of leaf components to support the reproductive
growth or due to some cue from the changing environmental conditions during the growing season. Field studies were conducted
in 1995–1996 at Stoneville, Mississippi, using six cotton genotypes and two planting dates (early and late), which produced
two distinctly different cotton populations reaching cutout at different times. Among the six genotypes were a photoperiod
sensitive line (non-flowering) and its counter part which had photoperiod insensitive genes backcrossed four times to the
photoperiod sensitive line (flowering). This pair was used to assess the degree that the photosynthetic decline could be attributed
to reproductive sink development. Leaf CO2-exchange rate (CER) and chlorophyll (Chl) fluorescence measurements were taken in mid-August, a period corresponding to cutout
for the early planted plots, and those leaves were collected. Leaf Chl level, soluble protein level, various soluble carbohydrate
levels and Rubisco activities were assayed on those leaves. Averaged across years, leaf CER and soluble protein levels were
reduced approximately 14% and 18%, respectively, for the early planted compared to the late planted cotton. Neither leaf Chl
levels or Chl fluorescence Fv/Fm values for Photosystem II yield were altered by the planting date. In 1996, leaves from the
non-flowering line had 12% greater Chl and 20% greater soluble protein levels than the flowering line. However, in 1996, the
CER of the early planted non-flowering line was reduced 10% compared to the late planted. Although remobilization of leaf
N to reproductive growth appears to be the principle component causing the cutout photosynthetic decline, the data also indicate
that environmental factors can play a small role in causing the decline.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
69.
Direct Measurement of Calcium Transport across Chloroplast
Inner-Envelope Vesicles 总被引:3,自引:1,他引:2
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Michael H. Roh Richard Shingles Michael J. Cleveland Richard E. McCarty 《Plant physiology》1998,118(4):1447-1454
The initial rate of Ca2+ movement across the inner-envelope membrane of pea (Pisum sativum L.) chloroplasts was directly measured by stopped-flow spectrofluorometry using membrane vesicles loaded with the Ca2+-sensitive fluorophore fura-2. Calibration of fura-2 fluorescence was achieved by combining a ratiometric method with Ca2+-selective minielectrodes to determine pCa values. The initial rate of Ca2+ influx in predominantly right-side-out inner-envelope membrane vesicles was greater than that in largely inside-out vesicles. Ca2+ movement was stimulated by an inwardly directed electrochemical proton gradient across the membrane vesicles, an effect that was diminished by the addition of valinomycin in the presence of K+. In addition, Ca2+ was shown to move across the membrane vesicles in the presence of a K+ diffusion potential gradient. The potential-stimulated rate of Ca2+ transport was slightly inhibited by diltiazem and greatly inhibited by ruthenium red. Other pharmacological agents such as LaCl3, verapamil, and nifedipine had little or no effect. These results indicate that Ca2+ transport across the chloroplast inner envelope can occur by a potential-stimulated uniport mechanism. 相似文献
70.
SSR markers closely associated with genes for resistance to root-knot nematode on chromosomes 11 and 14 of Upland cotton 总被引:2,自引:0,他引:2
Osman A. Gutiérrez Johnie N. Jenkins Jack C. McCarty Martin J. Wubben Russell W. Hayes Franklin E. Callahan 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2010,121(7):1323-1337
Molecular markers closely linked to genes that confer a high level of resistance to root-knot nematode (RKN) [Meloidogyne incognita (Kofoid & White) Chitwood] in cotton (Gossypium
hirsutum L.) germplasm derived from Auburn 623 RNR would greatly facilitate cotton breeding programs. Our objectives were to identify
simple sequence repeat (SSR) markers linked to RKN resistance quantitative trait loci (QTL) and map these markers to specific
chromosomes. We developed three recombinant inbred line (RIL) populations by single seed descent from the crosses of RKN-resistant
parents M-240 RNR (M240), developed from the Auburn 623 RNR source, moderately resistant Clevewilt 6 (CLW6), one of the parents
of Auburn 623 RNR, and susceptible parent Stoneville 213 (ST213). These crosses were CLW6 × ST213, M240 × CLW6, and M240 × ST213.
RILs from these populations were grown under greenhouse conditions, inoculated with RKN eggs, scored for root gall index,
eggs plant−1, and eggs g−1 root. Plants were also genotyped with SSR markers. Results indicated that a minimum of two major genes were involved in the
RKN resistance of M240. One gene was localized to chromosome 11 and linked to the marker CIR 316-201. This CIR 316-201 allele
was also present in CLW6 but not in Mexico Wild (MW) (PI593649), both of which are parents of Auburn 623 RNR. A second RKN
resistance gene was localized to the short arm of chromosome 14 and was linked to the SSR markers BNL3545-118 and BNL3661-185.
These two marker alleles were not present in CLW6 but were present in MW. Our data also suggest that the chromosome 11 resistance
QTL primarily affects root galling while the QTL on chromosome 14 mediates reduced RKN egg production. The SSRs identified
in this study should be useful to select plants with high levels of RKN resistance in segregating populations derived from
Auburn 623 RNR. 相似文献