Modern electronic and chemical thermometers used in the axilla are inaccurate

Rectal and axillary temperatures were measured simultaneously in 83 children using three different thermometer devices providing 166 pairs of results. In the first series consisting of 22 febrile children (44 measurements) and 20 afebrile children (40 measurements), the rectal mercury measurement was compared to an axillary mercury and axillary Tempa-DOT thermometer. The axillary mercury had sensitivity of 14/22 (64%) and specificity of 20/20 (100%) while the Tempa-DOT had sensitivity of 15/22 (68%) and specificity of 19/20 (95%). In the second series comprising 21 febrile children (42 measurements) and 20 afebrile children (40 measurements) the axillary mercury had sensitivity of 11/21 (52%) and specificity of 20/20 (100%) while the electronic thermometer had sensitivity of 10/21 (48%) and specificity of 20/20 (100%). Regardless of the thermometer used, the axilla is a poor alternative to rectal measurements in the diagnosis of fever. CONCLUSION: Mercury-free thermometers, when used in the axilla are as poor alternatives to rectal measurements as mercury-in-glass thermometers.     

Structure of very early protein folding intermediates: new insights through a variant of hydrogen exchange labelling

BACKGROUND: Hydrogen exchange labelling has been a key method in characterizing the structure of transient folding intermediates. In studies of several proteins, however, there has been clear spectroscopic evidence for partial folding of some kind at very early times, before any protection from exchange was measurable. These results, presumably a consequence of limited stability of specific backbone interactions, have made it difficult to assess the extent of native-like folding in the very early intermediates. We have used a variant of the labelling method to investigate marginally stable structures formed within the first few milliseconds of refolding of two such proteins, hen lysozyme and ubiquitin. RESULTS: In lysozyme, population of a subset of native-like secondary structures on this timescale is revealed, thus reconciling the exchange behaviour with circular dichroism measurements and confirming the significance of the rapidly formed embryonic structure as a foundation for the subsequent folding pathway. In the case of ubiquitin, by contrast, no significantly protective structure was detectable, suggesting that here secondary structural elements can be populated only marginally ahead of the major cooperative folding event; this was also supported by stopped-flow circular dichroism measurements. CONCLUSIONS: The hydrogen exchange approach can be extended to probe the formation of native-like structure formed in very early folding intermediates, even when the stability of specific interactions is marginal. In the case of lysozyme, this has provided a new window on an early stage of organization of the alpha-helical domain.     

Crystal structures of Escherichia coli and Salmonella typhimurium 3-isopropylmalate dehydrogenase and comparison with their thermophilic counterpart from Thermus thermophilus

The basis of protein stability has been investigated by the structural comparison of themophilic enzymes with their mesophilic counterparts. A number of characteristics have been found that can contribute to the stabilization of thermophilic proteins, but no one is uniquely capable of imparting thermostability. The crystal structure of 3-isopropylmalate dehydrogenase (IPMDH) from the mesophiles Escherichia coli and Salmonella typhimurium have been determined by the method of molecular replacement using the known structure of the homologous Thermus thermophilus enzyme. The structure of the E. coli enzyme was refined at a resolution of 2.1 A to an R-factor of 17.3%, that of the S. typhimurium enzyme at 1.7 A resolution to an R-factor of 19.8%. The three structures were compared to elucidate the basis of the higher thermostability of the T. thermophilus enzyme. A mutant that created a cavity in the hydrophobic core of the thermophilic enzyme was designed to investigate the importance of packing density for thermostability. The structure of this mutant was analyzed. The main stabilizing features in the thermophilic enzyme are an increased number of salt bridges, additional hydrogen bonds, a proportionately larger and more hydrophobic subunit interface, shortened N and C termini and a larger number of proline residues. The mutation in the hydrophobic core of T. thermophilus IPMDH resulted in a cavity of 32 A3, but no significant effect on the activity and thermostability of the mutant was observed.     

Modification of the calcium and calmodulin sensitivity of the type I adenylyl cyclase by mutagenesis of its calmodulin binding domain

The type I adenylyl cyclase is directly stimulated by Ca2+ and calmodulin in vitro, and the enzyme is also stimulated by increases in intracellular Ca2+ in vivo. Ca2+ stimulation of the enzyme in vivo may be due to direct interactions of the enzyme with Ca2+ and calmodulin or to an indirect mechanism involving stimulation of the enzyme by Ca(2+)-activated protein kinases. In this study, we have made several point mutations within the calmodulin binding domain to determine if the Ca2+ sensitivity of the enzyme can be modified by mutagenesis. The catalytic activities of the mutant enzymes were comparable to wild type type I adenylyl cyclase. Substitution of Cys-507 with Ser-507 did not have significant effects on the calmodulin or Ca2+ sensitivity of the enzyme. However, replacement of Lys-504 with Asp caused a 4-fold decrease in sensitivity to Ca2+. Ca2+ and calmodulin stimulation were abolished by substitution of Phe-503 with Arg-503. Stimulation of type I adenylyl cyclase activity in vivo by intracellular Ca2+ was also greatly diminished with the Arg-503 mutant indicating that Ca2+ stimulation of the enzyme in vivo is due primarily to direct interactions with calmodulin and Ca2+. These data demonstrate that the Ca2+ sensitivity of this enzyme can be modulated by point mutagenesis within the putative calmodulin binding domain and indicate that the enzyme can be directly regulated by Ca2+ and calmodulin in vivo.     

Fractures of the patella in children and adolescents

Renal scarring is considered a criterion of reflux nephropathy and the end stage of pyelonephritis. Prednisolone, a strong anti-inflammatory drug, at doses of 1 or 2 mg/kg prevented renal scarring in rats following infection with Serratia marcescens. Four or 8 mg/kg of prednisolone, however, did not inhibit renal scar formation. In a time course experiment, renal scarring was prevented when 4-day treatment with prednisolone was initiated 2, 5, or 13 days after infection. These results show that prednisolone is effective in preventing such scarring and suggest the clinical use of this drug for preventing renal scar formation after pyelonephritis and reflux nephropathy.