Specific techniques for assessing function: introduction.
Explore the source record for details and available documents.
Biomedical subjects
Publications and source records attributed to S Kaplan.
Explore the source record for details and available documents.
A biochemical analysis of the in vitro assembly of the form II ribulose-1,5-bisphosphate carboxylase/oxygenase from Rhodopseudomonas sphaeroides after transcription and translation from cloned DNA is presented. The predominant enzymatically active oligomeric forms of the in vitro-synthesized and -assembled ribulose-1,5-bisphosphate carboxylase are tetramers and hexamers. Assembly of the monomeric subunits to form active enzyme appears to be dependent on the presence of a minimum number of subunits in the cell extract. Assembly of ribulose-1,5-bisphosphate carboxylase also was observed when the protein-synthesizing extracts were prepared from cells which were partially derepressed for ribulose-1,5-bisphosphate carboxylase expression.
We report the cloning and characterization of the gene product of the gene for the form II ribulose bisphosphate carboxylase from Rhodopseudomonas sphaeroides. We present evidence that the form II enzyme is encoded by a single gene in R. sphaeroides; however, this gene does hybridize to a second chromosomal locus.
The mRNA levels specific for ribulose-1,5-bisphosphate carboxylase, light-harvesting I polypeptides alpha and beta, and reaction center polypeptides L and M were assayed by use of a series of DNA probes specific for each cognate mRNA. Both the steady-state amounts and sizes of the specific mRNAs were measured as a function of the light intensity incident to the culture, the presence or absence of oxygen, and the type of substrate present in the growth medium. Northern hybridization revealed at least two and possibly three transcripts for ribulose-1,5-bisphosphate carboxylase. The cellular level of mRNA specific for ribulose-1,5-bisphosphate carboxylase increased in consort with enzyme activity as a function of both light intensity and reducing state of the substrate. Neither mRNA nor enzyme activity was detectable in aerobically grown cells. For the light-harvesting I and reaction center polypeptides there exist two transcripts, the larger of which appears to be a polycistronic mRNA possessing information for all four polypeptides and a smaller transcript specific for only the alpha and beta polypeptides of the light-harvesting I complex. The regulation of each of these mRNAs was affected by light and oxygen, but was not significantly affected by the oxidation-reduction state of the substrate.
The cellular content of phospholipid transfer activity in Rhodopseudomonas sphaeroides was examined as a function of both oxygen partial pressure and light intensity used for growth. Cells grown under high light conditions (100 W/m2) had over two times the cellular level of phospholipid transfer activity when compared with cells grown under other conditions. Although cells grown under low light conditions (3 W/m2) had the lowest amount of total phospholipid transfer activity, they had the highest level (49%) of membrane-associated transfer activity. The soluble phospholipid transfer activity was further localized into periplasmic and cytoplasmic fractions. The distribution of phospholipid transfer activity in cells grown under medium light intensity (10 W/m2) was calculated as 15.1% membrane-associated, 32.4% in the periplasm, and 52.5% in the cytoplasm. The phospholipid transfer activities in the periplasmic and cytoplasmic fractions had distinctly different properties with respect to their molecular weights (56,000 versus 27,000) and specificities of transfer (phosphatidylethanolamine greater than phosphatidylglycerol versus phosphatidylglycerol greater than phosphatidylethanolamine).
We analyzed, by restriction endonuclease mapping and electron microscopy, the genome of the lytic Rhodopseudomonas sphaeroides-specific bacteriophage RS1 and characterized it as a linear molecule of approximately 60 to 65 kilobases. When the DNA from purified phage particles was examined by several independent methods, considerable size heterogeneity was apparent in the RS1 DNA. This size heterogeneity was concluded to be of biological origin, was independent of the specific host strain used to propagate virus, and was not due to the presence of host DNA within or nonspecifically associated with purified virions. In addition, treatment of RS1 DNA with either BAL 31 nuclease or DNA polymerase I Klenow fragment revealed that several distinct regions exist within the viral chromosome which contain free 3' hydroxyl groups. A restriction endonuclease map of the RS1 genome was constructed by using the restriction endonucleases EcoRI, ClaI, KpnI, BamHI, MluI, SmaI, and BclI; thereby allowing the positioning of some 40 restriction sites within the viral genome. The results are discussed in terms of the significance and the possible biological origin of the unique features discovered within the phage RS1 DNA.
A 7-year-old girl with incontinentia pigmenti and a history of multiple bacterial infections developed chronic meningitis with Actinomycosis odontolyticus, which was successfully treated with long-term penicillin administration. Although all tests of immunologic function were normal in this patient, her history of recurrent and unusual infections is consistent with previous suggestions of an undefined immunodeficiency state associated with incontinentia pigmenti.
An intervention was developed to increase patient involvement in care. Using a treatment algorithm as a guide, patients were helped to read their medical record and coached to ask questions and negotiate medical decisions with their physicians during a 20-minute session before their regularly scheduled visit. In a randomized controlled trial we compared this intervention with a standard educational session of equal length in a clinic for patients with ulcer disease. Six to eight weeks after the trial, patients in the experimental group reported fewer limitations in physical and role-related activities (p less than 0.05), preferred a more active role in medical decision-making, and were as satisfied with their care as the control group. Analysis of audiotapes of physician-patient interactions showed that patients in the experimental group were twice as effective as control patients in obtaining information from physicians (p less than 0.05). Results of the intervention included increased involvement in the interaction with the physician, fewer limitations imposed by the disease on patients' functional ability, and increased preference for active involvement in medical decision-making.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Although infants with neonatal polycythemia and hyperviscosity often present with cardiorespiratory distress, little information is available regarding the cardiac function of such babies before or after partial exchange transfusion. To assess cardiac function, we performed M-mode echocardiograms in 19 asymptomatic newborn infants (4 to 12 hours of age) who had venous hematocrits greater than 65%. The echocardiograms were performed immediately prior to and following partial exchange transfusion and were repeated at 48 hours of age. Eighteen matched newborn controls also underwent echocardiography within the first 12 hours of life and again at 48 hours. Polycythemic newborns had elevated right ventricular preejection period to right ventricular ejection time ratios compared with controls (0.46 +/- 0.11 v 0.37 +/- 0.04, P = .002), suggestive of increased pulmonary vascular resistance. These indexes normalized following partial exchange transfusion. In addition, the polycythemic newborns were relatively bradycardic prior to exchange (116 +/- 13 beats per minute v 125 +/- 16 beats per minute, P less than .05), but heart rates normalized following the procedure. At 48 hours, the polycythemic and control groups were different only in that the mean shortening fraction of the polycythemic group was lower than that of the controls (32% +/- 4% v 36% +/- 6%, P = .02). The findings are consistent with elevated pulmonary vascular resistance associated with polycythemia and hyperviscosity. Cardiac output may be lower in polycythemic infants. The finding of reduced fractional shortening in polycythemic infants at 48 hours following partial exchange transfusion is unexplained.
Explore the source record for details and available documents.
Structural changes association with the intracytoplasmic membrane during the cell cycle of the photosynthetic bacterium Rhodopseudomonas sphaeroides have been studied by freeze-fracture electron microscopy. The isolated intracytoplasmic membrane vesicles, chromatophores, were fused in order to obtain large fracture faces, allowing more precise measurements and statistical analysis of both intramembrane particle density and size determinations. The intramembrane particle density of the protoplasmic face (PF) of the intracytoplasmic membrane, (from 4970 to 8290/micrometers 2), was shown to be a linear function of the protein/phospholipid ratio (from 2.5 to 5.1, w/w) of the intracytoplasmic membrane. Under constant light intensity, both the average particle size and particle size distribution remained unchanged during the cell cycle. These results provide the structural basis for the earlier reported cell-cycle-specific variations in both protein/phospholipid ratio and alternation in phospholipid structure of the intracytoplasmic membrane of R. sphaeroides during photosynthetic growth. The average particle diameter in the PF face of the intracytoplasmic membrane was 8.25, 9.08 and 9.75 nm at incident light intensities of 4000, 500 and 30 ft X cd, respectively. When chromatophores were fused with small, unilamellar liposomes, the intramembrane particle density decreased as input liposome phospholipid increased, whereas the particle size remained constant and particle distribution became random.
A phospholipid transfer protein has been purified 280-fold from Rhodopseudomonas sphaeroides when compared to the 40-70% ammonium sulfate fraction derived from the crude cell supernatant in which the activity was originally found (Cohen, L.K., Lueking, D.R., and Kaplan, S. (1979) J. Biol. Chem. 254, 721-728). When compared to the crude cell lysate, the activity has been purified approximately 1,400-fold with a recovery of 12.5%. The active protein is a monomer with a molecular weight of 26,500, as estimated by sedimentation velocity and sedimentation equilibrium, and 27,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The protein can transfer all phospholipid species tested with the order of efficiency of transfer being phosphatidylglycerol greater than phosphatidylcholine greater than phosphatidylethanolamine.
Explore the source record for details and available documents.
Terminally ill cancer patients at a Veterans Administration hospital were randomly assigned to receive hospice or conventional care. The hospice care was provided both in a special inpatient unit and at home. 137 hospice patients and 110 control patients and their familial care givers (FCGs) were followed until the patient's death. No significant differences were noted between the patient groups in measures of pain, symptoms, activities of daily living, or affect. Hospice patients expressed more satisfaction with the care they received; and hospice patients' FCGs showed somewhat more satisfaction and less anxiety than did those of controls. Hospice care was not associated with a reduced use of hospital inpatient days or therapeutic procedures and was at least as expensive as conventional care.
The phospholipid biosynthetic enzyme activities: CDP-diglyceride synthetase, phosphatidylglycerophosphate synthetase, PGP phosphatase, phosphatidylserine (PS) synthase, PS decarboxylase, and S-adenosyl-L-methionine:phosphatidylethanolamine (AdoMet:PE) N-methyltransferase were detected in crude cell-free extracts of Rhodopseudomonas sphaeroides. CDP-diglyceride synthetase and phosphatidylglycerophosphate synthetase co-enriched with penicillin-binding protein activity, a known cytoplasmic membrane marker, throughout fractionation of cell-free extracts of both chemoheterotrophically and photoheterotrophically grown cells. PS decarboxylase also co-enriched with the cytoplasmic membranes in fractions derived from chemoheterotrophically and photoheterotrophically grown cells, but substantially greater quantities of PS decarboxylase activity was found in the chromatophores derived from photoheterotrophically grown cells than could be accounted for by cytoplasmic membrane contamination of this sample. PS synthase (60% of the recovered activity) and S-adenosyl-L-methionine:phosphatidylethanolamine N-methyltransferase (90% of the recovered activity) were found in the supernatant fraction after high speed centrifugation of crude cell lysates, suggesting that these enzyme activities were not tightly membrane associated. The localization of phospholipid biosynthetic enzyme activity in R. sphaeroides is discussed in terms of the biosynthesis of the photosynthetic membranes.
Paroxysmal supraventricular tachycardia developed in two infants with structurally normal hearts and no evidence of ventricular preexcitation. In both infants, the onset of the arrhythmia was associated with the presence of an indwelling jugular venous catheter in the body of the right atrium. Repeated episodes of the arrhythmia occurred while the catheter remained in place. Restoration and maintenance of sinus rhythm could only be achieved after withdrawal of the catheter.