The methylerythritol phosphate (MEP) pathway found in many bacteria governs the synthesis of isoprenoids, which are crucial lipid precursors for vital cell components such as ubiquinone. Because mammals synthesize isoprenoids via an alternate pathway, the bacterial MEP pathway is an attractive target for novel antibiotic development, necessitated by emerging antibiotic resistance as well as biodefense concerns. The first committed step in the MEP pathway is the reduction and isomerization of 1-deoxy-D-xylulose-5-phosphate (DXP) to methylerythritol phosphate (MEP), catalyzed by MEP synthase. To facilitate drug development, we cloned, expressed, purified, and characterized MEP synthase from Yersinia pestis. Enzyme assays indicate apparent kinetic constants of KM
DXP = 252 µM and KM
NADPH = 13 µM, IC50 values for fosmidomycin and FR900098 of 710 nM and 231 nM respectively, and Ki values for fosmidomycin and FR900098 of 251 nM and 101 nM respectively. To ascertain if the Y. pestis MEP synthase was amenable to a high-throughput screening campaign, the Z-factor was determined (0.9) then the purified enzyme was screened against a pilot scale library containing rationally designed fosmidomycin analogs and natural product extracts. Several hit molecules were obtained, most notably a natural product allosteric affector of MEP synthase and a rationally designed bisubstrate derivative of FR900098 (able to associate with both the NADPH and DXP binding sites in MEP synthase). It is particularly noteworthy that allosteric regulation of MEP synthase has not been described previously. Thus, our discovery implicates an alternative site (and new chemical space) for rational drug development.
One hundred seventy-nine obese patients (mean body mass index = 36.3) were retrospectively evaluated for the development of cholelithiasis associated with the use of a 2530-kJ/d (605-kcal) very-low-calorie diet (VLCD). Nine percent of patients had preexisting gallstones and 11% of patients developed gallstones either during or within 6 mo of completing the diet. Six percent had subsequent cholecystectomy. Ursodeoxycholic acid administered to one patient resulted in spontaneous stone dissolution whereas spontaneous dissolution occurred in three patients. Surveys of patients at three other programs using the same diet yielded similar incidence of gallstones. We conclude that rapid weight loss associated with the use of VLCD is associated with a significant incidence of gallstone formation. VLCD should be physician supervised because resolution of cholelithiasis spontaneously, with stone passage, or dissolution with ursodeoxycholic acid therapy may reduce the need for cholecystectomy.
Multiply By To obtain Length inch (in.) 2.54 centimeter (cm) foot (ft) 0.3048 meter (m) mile (mi) 1.609 kilometer (km) Area acre 0.004047 square kilometer (km 2) acre 0.001563 square mile (mi 2) square mile (mi 2) 2.590 square kilometer (km 2) Flow rate cubic foot per second (ft 3 /s) 0.02832 cubic meter per second (m 3 /s) cubic foot per second per square mile [(ft 3 /s)/mi 2 ] 0.01093 cubic meter per second per square kilometer [(m 3 /s)/km 2 ] Temperature in degrees Celsius (°C) may be converted to degrees Fahrenheit (°F) as follows:
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