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S M Lewis

Publications and source records attributed to S M Lewis.

At least 91 records · Page 5Linked to original sources
PubMed1993

P nucleotides in V(D)J recombination: a fine-structure analysis.

Antigen receptor genes acquire junctional inserts upon assembly from their component, germ line-encoded V, D, and J segments. Inserts are generally of random sequence, but a small number of V-D, D-J, or V-J junctions are exceptional. In such junctions, one or two added base pairs inversely repeat the sequence of the abutting germ line DNA. (For example, a gene segment ending AG might acquire an insert beginning with the residues CT upon joining). It has been proposed that the nonrandom residues, termed "P nucleotides," are a consequence of an obligatory end-modification step in V(D)J recombination. P insertion in normal, unselected V(D)J joining products, however, has not been rigorously established. Here, we use an experimentally manipulable system, isolated from immune selection of any kind, to examine the fine structure of V(D)J junctions formed in wild-type lymphoid cells. Our results, according to statistical tests, show the following, (i) The frequency of P insertion is influenced by the DNA sequence of the joined ends. (ii) P inserts may be longer than two residues in length. (iii) P inserts are associated with coding ends only. Additionally, a systematic survey of published P nucleotide data shows no evidence for variation in P insertion as a function of genetic locus and ontogeny. Together, these analyses establish the generality of the P nucleotide pattern within inserts but do not fully support previous conjectures as to their origin and centrality in the joining reaction.

Animals↗
PubMed1993

Comparison of gavage, water bottle, and a high-moisture diet bolus as dosing methods for quantitative D-xylose administration to B6D2F1 (Mus musculus) mice.

Gavage, water bottle, and diet incorporation are 3 dosing methods used orally to administer test compounds to rodents. These 3 methods were compared in mice to determine which represented the most quantitative delivery system. For dietary incorporation, a high-moisture bolus form of NIH-31 rodent meal was developed using hydroxypropyl methylcellulose as an autoclave-stable binding agent. A high-moisture bolus was selected to increase the acceptability of the dosed diet and to promote quantitative consumption through reduced wastage. The test compound used was D-xylose, a pentose sugar that may be quantitatively detected, colorimetrically, in urine following oral dosing. Six male and 6 female B6D2F1 mice were placed in metabolism cages and dosed with a known quantity of D-xylose by each of the 3 methods. Urine was collected before and after each method of administration and analysed for total D-xylose; the per cent recovery was based upon the amount of D-xylose consumed. Quantitative consumption was apparently greatest for water bottle dosing with an average recovery of 56.0% of the original D-xylose dose. High-moisture bolus incorporation ranked second with 50.0% D-xylose recovery, and gavage was third with 41.0% D-xylose recovery.

Animal Feed↗
PubMed1992

Resolved conformational states of spin-labeled Ca-ATPase during the enzymatic cycle.

We have used frequency- and time-resolved electron paramagnetic resonance (EPR) to study the effects of substrate on the nanosecond conformational dynamics of the Ca-ATPase of sarcoplasmic reticulum, as detected by an iodoacetamide spin label (IASL) attached covalently to the enzyme. We confirm previous results [Coan, C. (1983) Biochemistry 22, 5826] showing that this probe is less rotationally mobile following the addition of nucleotides (ADP, AMPPNP, ATP) and that the shape of the spectrum suggests the presence of two components. We used two approaches to enhance EPR resolution in order to resolve the spectral components and their corresponding conformational states. First, to improve resolution in the frequency (spectral) domain, we used perdeuterated IASL, which results in narrower line widths. Digital spectral analysis resolves the EPR spectrum into two components, one that is indistinguishable from the spectrum observed in the absence of ligands and another that indicates more restricted probe motions, suggesting a distinct conformation of the labeled protein. Additions of substrate ligands appear to change only the mole fractions of the two components. The mole fraction of the restricted component (fR) was 0 in the absence of ligands, but increased to about 0.5 in the presence of saturating concentrations of AMPPNP and Ca2+. In general, ATP and its analogs increase fR, with larger effects observed in the presence of Ca. However, calcium has no effect by itself (fR = 0). Both monovanadate and decavanadate increase fR, but the formation of a covalent phosphoenzyme from inorganic phosphate (E2-P) had no effect (fR = 0).(ABSTRACT TRUNCATED AT 250 WORDS)

Adenine Nucleotides↗
PubMed1992

Oxidative DNA damage levels in rats fed low-fat, high-fat, or calorie-restricted diets.

Increased fat and caloric content of the diet has been associated with increased mammary tumor incidence. The dietary modulation of cellular redox state may be one mechanism behind this association. We have examined the effects of changes in dietary fat and caloric intake on the levels of 5-hydroxymethyluracil in DNA from rat liver and mammary gland. Female Fischer 344 rats, 40 days old, were maintained on 3% (low-fat), 5% (control), or 20% (high-fat) corn oil diets for 2 weeks. A fourth group of rats had the same daily fat intake as the control group, but total caloric intake was restricted by 40%. As a measure of oxidative DNA damage, 5-hydroxymethyluracil levels were measured in the DNA extracted from liver and mammary gland by gas chromatography-mass spectrometry. 5-Hydroxymethyluracil levels in the liver DNA of the low-fat, high-fat, and calorie-restricted groups were decreased relative to that of control, but the only significant decrease was in the calorie-restricted group (p less than 0.01). In the mammary gland DNA, statistically significant decreases in damage were found in each group relative to control (p less than 0.05). The relationship between fat in the diet and oxidative stress is thus complex. These results show that changes in dietary intake of both fat and calories can modulate oxidative DNA damage levels, and the effect of diet was more clearly evident in the DNA from mammary gland than in DNA from liver.

Animals↗
PubMed1992

The selection of laboratory equipment.

The last few years have seen dramatic changes in laboratory haematology. Technological developments with increasing levels of automation have resulted in a plethora of new instruments. The choice of which to buy, often confused by conflicting advice and persuasive salesmanship, must be balanced by the need for cost containment. This calls for an understanding of the principles of selection to ensure that the system that is chosen will provide reliable results in the most efficient manner at the lowest possible cost. Most of the instruments which are currently available perform their intended tasks reasonably well albeit with some inter-instrument differences, usually due to different operating procedures and different methods of calibration. The task is to find a system that performs the required tests correctly and reliably, fits into the work requirements of the laboratory and is accepted by the staff who will operate it, whilst its cost both capital and recurrent, should be within the department's financial constraints. The selection of laboratory equipment is thus a complex and time consuming task.

Cost-Benefit Analysis↗
PubMed1992

The New Jersey health care delivery structure--a clarification.

Recent SBME rules were developed from issues raised by a previous commissioner of health. SBME received comments on these issues at two public hearings. SBME drafted the rules to incorporate the substance and the principles of the state's Health Care Cost Reduction Act.

Coroners and Medical Examiners↗
PubMed1992

Single-operator comparison of early and mid-second-trimester amniocentesis.

We sought to determine whether early amniocentesis is a safe and acceptable method of genetic evaluation in early pregnancy. During the 54-month period from September 1986 to February 1991, 300 consecutive early second-trimester amniocenteses were performed transabdominally at 13-14 weeks' gestation and 567 consecutive mid-second-trimester transabdominal amniocenteses were performed at 16-18 weeks. Group assignment was nonrandomized, interoperator-dependent variables were eliminated, and analysis was performed in one cytogenetics laboratory. The median maternal age and indications for the procedure were similar in both groups. There were no significant differences between the early- and mid-second-trimester amniocenteses in failed sampling, ambiguous results, pregnancy loss from 4 weeks after the procedure to 28 weeks' gestation, preterm birth, or perinatal death rate. Pregnancy loss within 4 weeks of amniocentesis was more frequent in early- than in mid-second-trimester amniocenteses. We conclude that early amniocentesis is a safe and acceptable method of genetic evaluation.

Adult↗
PubMed1991

Microsecond rotational dynamics of spin-labeled Ca-ATPase during enzymatic cycling initiated by photolysis of caged ATP.

We have measured the microsecond rotational motions of the sarcoplasmic reticulum (SR) Ca-ATPase as a function of enzyme-specific ligands, including those that induce active calcium transport. We labeled the Ca-ATPase with a maleimide spin probe and detected rotational dynamics using saturation-transfer electron paramagnetic resonance (ST-EPR). This probe's ST-EPR spectra have been shown to be sensitive to microsecond protein rotational motion, corresponding to large-scale protein rotations that should be affected by changes in the enzyme's shape, flexibility, protein-protein interactions (oligomeric state), and protein-lipid interactions. We found that the motions of the enzyme-nucleotide and the enzyme-nucleotide/Ca states are indistinguishable from the motions in the absence of ligands. Rotational mobility does decrease in response to the addition of DMSO, a solvent that inhibits Ca-ATPase activity and stabilizes the phosphoenzyme. However, the addition of phosphate to form phosphoenzyme, in the presence or absence of DMSO, does not change the motions significantly. During the steady state of active calcium transport, the microsecond rotational mobility is indistinguishable from that of the resting enzyme. In order to detect any transient changes in mobility that might not be detectable in the steady state and to improve the precision of steady-state measurements, we photolyzed caged ATP with a laser pulse in the presence of calcium and detected the ST-EPR response from the spin-labeled enzyme, with a time resolution of 1 s.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1991

Cutting and closing without recombination in V(D)J joining.

Open and shut junctions are rare (V(D)J joining products in which site-specific recognition, cleavage and re-ligation of joining signals has been uncoupled from recombination. Here, we investigate the relationship of opening and shutting to recombination in two ways. First, we have tested a series of substrates containing one or two joining signals in an in vivo assay. Opening and shutting can be readily observed in substrates that have only one consensus joining signal. Thus, unlike recombination, the majority of open and shut events do not require interactions between two canonical joining signals. Next we examined two-signal substrates to investigate the effect of signal proximity on the frequency of dual open and shut events. These experiments indicate that at least some of the time opening and shutting can be a two-signal transaction. Together these results point to two mechanistically related, but distinct origins for open and shut joining events. In one case, cutting and closing may occur without interaction between two signals. In the other, we suggest that interaction of a canonical signal with 'cryptic' signal-like elements whose sequence is extensively diverged from canonical signals, may bias the V(D)J recombination machinery towards opening and shutting rather than recombination. Open and shut operations could in this way provide a means whereby mistakes in target recognition by the V(D)J recombination machinery produce a non-recombinant outcome, avoiding deleterious chromosomal rearrangements in lymphoid tissues.

Animals↗
PubMed1991

Primary thrombocythaemia in pregnancy.

Nine pregnancies in six patients with primary thrombocythaemia are reported. Eight pregnancies resulted in the delivery of normal infants. One pregnancy ended in spontaneous abortion at 7 weeks gestation. One pregnancy was complicated by superficial thrombophlebitis and a postpartum haemorrhage. We suggest that pregnancy in patients with primary thrombocythaemia can have a favourable outcome, but requires close monitoring. Administration of aspirin during pregnancy may be of benefit.

Abortion, Incomplete↗
PubMed1991

Lauryl sulphate haemoglobin: a non-hazardous substitute for HiCN in haemoglobinometry.

The haemiglobincyanide (HiCN) method for measuring haemoglobin is used extensively worldwide; its advantage is the ready availability of a stable and internationally accepted reference standard/calibrator. However, its use may create a problem, especially in automated analysers, as the waste disposal of large volumes of reagent containing cyanide may constitute a potential toxic hazard. As an alternative, conversion of haemoglobin to a sulphate derivative by nontoxic sodium lauryl sulphate has been proposed and is available as a commercial product from Toa Medical Electronics Co. Ltd., Japan. This evaluation has shown it to be as reliable and reproducible as HiCN for measuring haemoglobin at all concentrations, by both spectrophotometry and automated analyser. Maximum absorbance is at 534 nm. Total conversion occurs almost instantaneously, and includes methaemoglobin but not sulphaemoglobin. HbF is also measured. The only disadvantage is that solutions of haemoglobin-sulphate are stable for only a few hours at room temperature and up to three weeks at 4 degrees C. Thus, for standardization and calibration of instruments it is necessary to use a sample of blood or lysate the haemoglobin value of which has been assigned initially by the HiCN method.

Calibration↗