Dramatic differences in the binding of UDP-galactose and UDP-glucose to UDP-galactose 4-epimerase from Escherichia coli

UDP-galactose 4-epimerase catalyzes the interconversion of UDP-galactose and UDP-glucose during normal galactose metabolism. Within recent years the enzyme from Escherichia coli has been studied extensively by both biochemical and X-ray crystallographic techniques. One of several key features in the catalytic mechanism of the enzyme involves the putative rotation of a 4'-ketopyranose intermediate within the active site region. The mode of binding of UDP-glucose to epimerase is well understood on the basis of previous high-resolution X-ray crystallographic investigations from this laboratory with an enzyme/NADH/UDP-glucose abortive complex. Attempts to prepare an enzyme/NADH/UDP-galactose abortive complex always failed, however, in that UDP-glucose rather than UDP-galactose was observed binding in the active site. In an effort to prepare an abortive complex with UDP-galactose, a site-directed mutant protein was constructed in which Ser 124 and Tyr 149, known to play critical roles in catalysis, were substituted with alanine and phenylalanine residues, respectively. With this double mutant it was possible to crystallize and solve the three-dimensional structures of reduced epimerase in the presence of UDP-glucose or UDP-galactose to high resolution. This study represents the first direct observation of UDP-galactose binding to epimerase and lends strong structural support for a catalytic mechanism in which there is free rotation of a 4'-ketopyranose intermediate within the active site cleft of the enzyme.     

Cyclosporine dosage can be reduced when used in combination with an anti-intercellular adhesion molecule-1 monoclonal antibody in rats undergoing heterotopic heart transplantation

OBJECTIVE: Although occupationally embedded exercise is becoming a recognized topic in occupational therapy research, study of this phenomenon in children has been limited. The present study examined whether play elicits therapeutic patterns of movement through the use of two ABA single-case experiments. It was predicted that in two children with hypotonic cerebral palsy, the addition of a favorite game to the occupational form, or treatment environment, would increase functional vertical neck and back extension. METHOD: Each subject was positioned in prone on a wedge. During the A phases, each was given verbal directions to hold her head up. During the B phase, a favorite game was introduced. It was predicted that each subject would extend her back and neck in a functional way in order to manipulate and observe the game. The dependent variables were the mean range of neck extension and the mean range of back extension, which were measured by a videotaped, two-dimensional kinematic analysis. RESULTS: The addition of a favorite game to the occupational form improved vertical neck and back extension while discouraging nonfunctional neck hyperextension and fixing postures in both children. CONCLUSION: Embedding exercise within a play occupation enhanced the prone extension of two children with hypotonic cerebral palsy.     

Solar disinfection of drinking water contained in transparent plastic bottles: characterizing the bacterial inactivation process

A series of experiments is reported to identify and characterize the inactivation process in operation when drinking water, heavily contaminated with a Kenyan isolate of Escherichia coli, is stored in transparent plastic bottles that are then exposed to sunlight. The roles of optical and thermal inactivation mechanisms are studied in detail by simulating conditions of optical irradiance, water turbidity and temperature, which were recorded during a series of solar disinfection measurements carried out in the Kenyan Rift Valley. Optical inactivation effects are observed even in highly turbid water (200 ntu) and at low irradiances of only 10 mW cm-2. Thermal inactivation is found to be important only at water temperatures above 45 degrees C, at which point strong synergy between optical and thermal inactivation processes is observed. The results confirm that, where strong sunshine is available, solar disinfection of drinking water is an effective, low cost method for improving water quality and may be of particular use to refugee camps in disaster areas. Strategies for improving bacterial inactivation are discussed.     

Minimally invasive surgery for coronary disease: a new alternative

To avoid the inflammatory syndrome generated by cardiopulmonary bypass, a new surgical technique, minimal invasive direct coronary artery bypass (MIDCAB), has been developed. An anastomosis is performed between the left internal mammary artery (LIMA) and the left anterior descending artery (LAD) on a beating heart, through a limited anterior thoracotomy. We describe our experience with this technique. Ten consecutive patients underwent a MIDCAB procedure. (9 males, age 65.9 +/- 9 years). There were 8 bypasses of the LIMA on the LAD, one bilateral mammary bypass on the LAD and the right coronary artery, and one conversion to a standard sternotomy with CPB for a saphenous vein bypass on the LAD because of injury to the LIMA (2nd case). There was one redo for haemostasis of the mammary artery bed (3rd case). The first 3 patients required postoperative blood transfusion. From the 4th operation onwards, with the introduction of new instrumentation which was better adapted to the narrowness of the surgical field, there were no further surgical complications. During the follow-up (mean 5 months; range 2-9), no patient suffered anginal recurrence. With the improvement of instrumentation, the MIDCAB technique offers satisfactory short- and mid-term results, while avoiding CPB with its adverse effects. Lastly, the cosmetic result is far better than with the conventional procedure.     

Prevalence and management of major risk factors in 500 men with stable coronary disease

In order to achieve an overview of neuroradiology in Europe a questionnaire was sent to all ESNR National Delegates. The answers received were submitted to a data-based analysis, leading to the conclusion that neuroradiology is an expanding discipline among neurological sciences.     

Lipoprotein nature of Bacillus licheniformis membrane penicillinase

Membrane penicillinase (penicillin amido-beta-lactamhydrolase, EC 3.5.2.6) from Bacillus licheniformis bears a striking resemblance to the major outer membrane lipoprotein of Escherichia coli. It can be specifically labeled in vivo with [3H]glycerol, [35S]cysteine, or [3H]palmitate but not by [32P]orthophosphate. The labeled residues are located at or near the NH2 terminus of the membrane penicillinase because they can be completely removed by trypsin which cleaves a hydrophobic peptide(s) from the NH2 terminus, thereby rendering the enzyme hydrophilic. The membrane penicillinase produced by the 749/C gene carried in E. coli on phage lambda is similar to the enzyme formed in strain 749/C itself. The peptide antibiotic globomycin, which prevents processing of the E. coli prolipoprotein, severely inhibited the attachment of [3H]palmitate or [3H]glycerol to the 749/C enzyme (either in B. licheniformis 749/C or in E. coli), blocked the accumulation of penicillinase in the plasma membrane, and enhanced the formation of exoenzyme. Under the same conditions, globomycin does not prevent the attachment of palmitate or glycerol to the E. coli prolipoprotein but inhibits processing of the modified precursor to the mature lipoprotein. These results are in contrast with the lack of effect of globomycin on the RTEM-beta-lactamase of E. coli which has no detectable hydrophobic membrane form and was not labeled with palmitate or glycerol.