Inhibition of photosystem II by uncouplers at alkaline pH and its reversal by artificial electron donors.

When chloroplasts are aged for 5 min at pH 9.6, or are exposed to uncouplers at pH 8.5-9.0, electron flow from water to Hill acceptors is inhibited. Both treatments induce rapid millisecond dark decay of delayed light emission. 3-(3,4-Dichlorophenyl)-1,1-dimethylurea-sensitive electron transport through Photosystem II can be regenerated in both types of inhibited chloroplasts by the artificial electron donor, 1, 5-diphenylcarbohydrazide. Neither treatment inhibits electron flow through Photosystem I. Uncouplers at alkaline pH, when added in the light, are less effective in producing the inhibition than when added in the dark. These results are interpreted as indicating inhibition of the oxygen-evolving apparatus by alkaline intrathylakoid pH.

The totally occluded internal carotid artery. Preliminary observations using rapid sequential computerized tomographic scanning.

Cerebral angiography often cannot distinguish between complete thrombosis or fibrosis of the internal carotid artery (ICA) and nonvisualization due to a total occlusion of the common carotid or origin of the ICA. Whereas surgery may be beneficial if the distal carotid is patent (type 1), thromboendarterectomy may be contraindicated if thrombus or fibrosis extend to the intracranial branches (type 2). Rapid sequential computerized tomography (RSCT) was used to examine 15 patients whose ICAs appeared occluded by angiography. Of four ICAs classed as type 1 by RSCT, three were found to be patent during surgical exploration, and carotid reconstruction was successfully performed. Three other ICAs classed as type 2 by RSCT were also surgically explored, and complete thrombosis was confirmed. The RSCT technique provides an effective and nonoperative means of determining whether a nonvisualized ICA is reconstructible.

Economies of scope in physicians' work: the performance of multiple surgery.

While the fees for the large majority of physicians' services in the new Medicare Fee Schedule (MFS) are derived directly from studies of the resource costs involved, payments for one class of procedures, multiple surgery, are based instead on existing policies and conventions. Using surveys of physicians, we measured the work and time involved in performing 146 multiple surgeries. We found economies of scope exist in performing these services, particularly during the preoperative and postoperative periods. We also found some differences in economies across procedures. Based on our findings, we propose payment policies for multiple surgery.

Characteristics of the Mg-ATPase Activity Associated with the Membrane-Bound Maize Coupling Factor.

The membrane-bound coupling factor of maize mesophyll thylakoids is a latent ATPase. Mg(2+)-ATPase activity can be induced in the light with either dithiothreitol or low concentrations of trypsin. Maize thylakoids that are activated with light plus trypsin exhibit considerably higher levels of activity in Na(2)SO(3)-dependent Mg(2+)-ATPase assays compared to thylakoids that are light and dithiothreitol activated (1400 micromoles per milligram of chlorophyll per hour versus 200 micromoles per milligram of chlorophyll per hour). Treatment with light and dithiothreitol or light plus trypsin were also required to demonstrate high levels of octyl glucoside-dependent Mg(2+)-ATPase activity in maize mesophyll thylakoids. Only small differences in octyl glucoside-dependent Mg(2+)-ATPase activity were observed in preparations that were activated in the light with either trypsin or dithiothreitol. Mg(2+)-ATPase activity can also be induced in maize mesophyll chloroplasts by illuminating intact preparations under appropriate conditions. Little or no ATPase activity was observed in the absence of illumination or in the presence of light plus methyl viologen. The active state decayed in the dark with a t((1/2)) of 6 to 7 minutes at room temperature. Based on the effect of the thiol oxidant, o-iodosobenzoate, and the uncoupler, nigericin, on the kinetics of deactivation of ATPase activity in intact maize chloroplasts, it appears that the activation process requires a transmembrane proton gradient and reduction of a key disulfide bridge in the gamma of chloroplast coupling factor one.

A proposed mechanism for the stimulatory effect of bicarbonate ions on ATP synthesis in isolated chloroplasts.

The effect of bicarbonate ions on induction of Mg(2+)-ATPase activity, on the N-ethylmaleimide inhibition of phosphorylation and on energy-dependent adenine nucleotide exchange has been examined with pea seedling chloroplasts. Incubation of chloroplasts with N-ethylmaleimide in the presence of 15 millimolar bicarbonate in the light results in enhanced inhibition of ATP synthesis when the preillumination pH is maintained between 7.0 and 7.5. Bicarbonate also enhances Mg(2+)-ATPase activity when it is included in the light-triggering stage at pH 7.0. The conditions (medium pH, bicarbonate concentration, etc.) for demonstrating the bicarbonate-induced enhancement of the N-ethylmaleimide inhibition and ATPase activity are similar to those required for the direct effect of bicarbonate on phosphorylation. Bicarbonate, under the same conditions, does not affect adenine nucleotide exchange (binding or release). It is concluded that the stimulatory effect of bicarbonate on ATP synthesis may be related to its ability to alter directly the conformation of the chloroplast coupling factor under conditions (suboptimal pH) where the enzyme shows minimal activity.

Further studies on the bicarbonate stimulation of photophosphorylation in isolated chloroplasts.

The bicarbonate effect in stimulating the rate of photophosphorylation by isolated spinach (Spinacia oleracea var. Virginia blight-resistant savoy) chloroplasts at a pH below the optimum has been re-examined. Its seasonal nature may be related to the hormonal status of the plants. Bicarbonate anions stimulate adenosine 5'-triphosphate synthesis if added in the final, adenosine 5'-triphosphate-forming stage of either a postillumination or an acid-base experiment. They also stimulate the membrane-bound, Mg(2+)-dependent adenosine 5'-triphosphatase of chloroplasts, and the Ca(2+)-dependent adenosine 5'-triphosphatase of detached coupling factor. These and other data point to the interaction between energized thylakoid membranes and the coupling factor as the probable site of action of bicarbonate anions when they stimulate photophosphorylation.

Sulfhydryl reagents and energy-linked reactions in monocot thylakoids.

Monofunctional maleimides have been used to covalently modify the coupling factor protein of monocot thylakoid membranes. As with dicot thylakoids, incubation of the monocot thylakoids with maleimides in the light but not in the dark results in inhibition of both ATP synthesis and hydrolysis. In the dark, sites on the gamma and epsilon subunits of maize Zea mays coupling factor 1 are modified after incubation of maize mesophyll thylakoids with the fluorescent maleimide N-(anilinonaphthyl-4) maleimide. A light accessible site localized solely to the gamma subunit has also been demonstrated. In contrast to the case with dicot thylakoids (spinach [Spinacia oleracea] and pea [Pisum sativum]) treatment of monocot thylakoids (maize, barley [Hordeum vulgare], crabgrass [Digitaria sanguinalis]) with bifunctional maleimides or thiol oxidants in the light does not result in functional uncoupling, i.e the bifunctional reagents act more like energy transfer inhibitors. The lack of functional uncoupling could be due either to a failure of the reagents to cross-link key sulfhydryl residues in the gamma subunit or to the continued ability of the gamma subunit to gate proton movements through the chloroplast coupling factor complex even though its conformation has been altered by sulfhydryl reagents.

The resource-based relative value scale: methods, results and impacts on urology.

Increased concern for rising health care costs in the United States has led to the passage of legislation to reform physician payment for Medicare services based on resource inputs. In January 1992 the Health Care Financing Administration began implementing the new law, which replaces the existing Medicare system of physician payment with a fee schedule based on the resource-based relative value scale (RBRVS). We summarize the methods and data used to derive the RBRVS for urology. A national random sample of 115 practicing urologists completed structured telephone surveys to provide ratings of physician time and work required before, during and after most frequently performed urological services. Subsequent survey cycles with urologists provided further refinement. Urologists then participated in a cross-specialty physician panel to link services from all specialties onto a common scale. This common scale was adjusted for geographic differences in practice overhead costs and malpractice insurance premiums. A monetary conversion factor, determined by the Health Care Financing Administration, was then applied to convert the RBRVS into a Medicare fee schedule. The merits and demerits of the scientific process used to develop and maintain the relative value scale are extensive. While statistically valid and reproducible, the study results have been altered in the political arena. The results and impacts of the new Medicare payment system on urology will be significant, although it is not yet clear how urological practice will be affected. Although faring better than most surgical specialties, urologists stand to lose approximately 8% of their Medicare income when the new fee schedule is fully implemented. There will be relative gains for evaluation and management services and losses for most invasive procedures.

The Coupling of Electron Flow to ATP Synthesis in Pea and Maize Mesophyll Chloroplasts : I. INTERACTION OF ADENINE NUCLEOTIDES AND ENERGY TRANSFER INHIBITORS WITH THE COUPLING FACTOR COMPLEX.

The rate of nonphosphorylating electron transport (in the absence of ADP and inorganic phosphate) in well-coupled (ATP/2e(-) = 0.9-1.1) maize mesophyll chloroplasts is not modulated by external pH (6.5-8.5), low levels of ADP or ATP, or energy transfer inhibitors, e.g. triphenyltin and Hg(2+) ions. In contrast nonphosphorylating electron flow in pea chloroplasts is sensitive to alterations in medium pH, and to the presence of adenine nucleotides and energy transfer inhibitors in the assay medium. Although ATP is without effect on the rate of basal electron transport in maize chloroplasts, steady-state proton uptake is stimulated 3- to 5-fold by low levels of ATP. These results suggest that differences may exist in the manner in which the coupling factor complex controls proton efflux from the intrathylakoid space in C(3) and C(4) mesophyll chloroplasts.

l-Arginine and l-Canavanine Metabolism in Jack Bean, Canavalia ensiformis (L.) DC. and Soybean, Glycine max (L.) Merr.

Studies have been conducted with the arginase (l-arginine amidinohydrolase, EC 3.5.3.1) of two legumes: jack bean, Canavalia ensiformis (L.) DC., a l-canavanine-containing plant and soybean, Glycine max, a canavanine-free species. Analyses of the arginase obtained from gradient-purified mitochondria of these legumes revealed that the arginine-dependent (ADA) and canavanine-dependent activities (CDA) were localized within this organelle.Kinetic analyses of affinity-purified mitochondrial arginase revealed an apparent K(m) of 7 to 8 millimolar for arginine with both the jack bean and soybean arginases. Comparable determinations with canavanine revealed an apparent K(m) of 38 millimolar with the jack bean enzyme; the affinity for this arginine analog with the soybean enzyme is so poor that product formation remained linear even with a canavanine concentration of 890 millimolar.A single macromolecule appears to be responsible for both the ADA and CDA of jack bean arginase. Ion-exchange chromatography of mitochondrial arginase revealed that the ADA and CDA eluted as a single, discrete peak from DEAE-cellulose. Analyses with arginine- and canavanine-linked Sepharose failed to reveal more than one enzyme. Both the ADA and CDA increased by nearly identical amounts following elution from arginine- and canavanine-linked cyanogen bromide-activated sepharose. Neither ADA nor CDA increased preferentially over the other.

Interaction of 2-n-heptyl-4-hydroxyquinoline-N-oxide with photosystem II in chloroplasts and subchloroplast particles.

The effects of 2-n-heptyl-4-hydroxyquinoline-N-oxide on electron transport in thylakoids and oxygen-evolving photosystem II particles has been examined. Kinetic fluorescence studies reveal that the site of inhibition for alkyl derivatives of hydroxyquinoline-N-oxide (I50 approximately equal to 2 microM) is located between Q and plastoquinone. Studies with thylakoids isolated from atrazine-resistant pigweed plants indicate that the modification in the Q/B membrane complex that confers increased resistance to inhibition by atrazine also results in decreased sensitivity to inhibition by 2-n-heptyl-4-hydroxyquinoline-N-oxide (resistant/ sensitive ratio = 11). From the results of tetramethylphenylenediamine by-pass experiments, determinations of inhibitor sensitivity in trypsin-treated thylakoids and competitive displacement experiments made with [14C]metribuzin in thylakoids and photosystem II particles, it is suggested that 2-n-heptyl-4-hydroxyquinoline-N-oxide binds in a region of the Q/B complex that is distinct from the 3-(3,4-dichloro)-1,1-dimethyl urea and atrazine binding sites.

A small genetic region that controls dihydroorotate dehydrogenase in Drosophila melanogaster.

A locus is described that controls levels of mitochondrial dihydroorotate dehydrogenase (EC 1.3.3.1) in Drosophila melanogaster. The effects of alleles of the locus, Dhod, are manifest in preparations from whole organisms as well as in partially purified mitochondrial preparations; however, other mitochondrial functions do not appear to be appreciably affected by Dhod genotypes. The locus maps near p in the proximal portion of the right arm of chromosome 3. Flies trisomic for a chromosome segment including that region display elevated enzyme levels, implying that an enzyme structural gene is in that vicinity. Furthermore, Dhod alleles are semidominant in heterozygotes, suggesting that the dosage-sensitive element detected in the trisomics is actually the Dhod locus. These findings are discussed relative to the role of dihydroorotate dehydrogenase in the de novo pyrimidine biosynthetic pathway and relative to other pathway mutants that have been described in Drosophila.

Inhibition of photosynthetic electron transport by diphenyl ether herbicides.

The effects of the diphenyl ether herbicides HOE 29152 (methyl-2[4-(4-trifluoromethoxy) phenoxy] propanoate) and nitrofluorfen (2-chloro-1-[4-nitrophenoxy]-4-[trifluoromethyl]benzene) on photosynthetic electron transport have been examined with pea seedling and spinach chloroplasts. Linear electron transport (water to ferricyanide or methylviologen) is inhibited in treated chloroplasts, but neither photosystem II activity (water to dimethylquinone plus dibromothymoquinone) nor photosystem I activity (diaminodurene to methylviologen) is affected. Cyclic electron flow, cata-lyzed by either phenazine methosulfate or diaminodurene, is resistant to inhibition by nitrofluorfen. In diphenyl ether-treated chloroplasts the half-time for the dark reduction of cytochrome f is increased 5- to 15-fold. These data indicate that the site of inhibition for the diphenyl ethers is between the two photosystems in the plastoquinone-cytochrome f region.