No abstract

The apparent instability of ,B-galactosidase in toluene-treated cells or cell-free extracts of lactic streptococci is explained by the fact that these organisms do not contain the expected enzyme. Instead, various strains of Streptococcus lactis, S. cremoris, and S. diacetilactis were shown to hydrolyze o-nitrophenyl-f3-D-galactoside-6-phosphate (ONPG-6-P), indicating the presence of a different enzyme. In addition, lactose metabolism in S. lactis C2F was found to involve enzyme I (El), enzyme II (EII), factor III (FIII), and a heat-stable protein (HPr) of a phosphoenolpyruvate (PEP)-dependent phosphotransferase system analogous to that of Staphylococcus aureus. Mutants of S. lactis C2F, defective in lactose metabolism, possessed the phenotype lac-gal-. These strains were unable to accumulate 4{Cthiomethyl-,3-D-galactoside, to hydrolyze ONPG, or to utilize lactose -when grown in lactose or galactose broth. In addition, these mutants contained EI and HPr, but lacked EII, FIII, and the ability to hydrolyze ONPG-6-P. This suggested that the defect was in the phosphorylation step. Lactose-negative mutants of S. lactis 7962, a strain containing j3-galactosidase, could be separated into several classes, which indicated that this organism is not dependent upon the PEP-phosphotransferase system for lactose metabolism.on August 4, 2020 by guest

Objective: To compare ultrasound (US)-guided vs. landmark-guided techniques for central venous access (CVA) in the emergency department. Methods: This was a prospective study of consecutive patients enrolled at a university teaching hospital with an annual census of approximately 100,000. On even days patients had CVA with ultrasonic assistance; patients presenting on odd days had CVA via traditional landmark techniques. Ultrasound users were emergency medicine faculty or residents who completed a one-hour training session. A data collection tool with 17 variables was completed for each central line placed. Variables were compared using the independent t-test, Fisher's exact test, and the nonparametric Mann-Whitney U test. Results: Between August 1, 2000, and February 1, 2001, data for 122 subjects (n = 51 for US, and n = 71 for landmark) were collected. Variables with statistically significant differences are as follows. Mean (ϮSD) time from skin puncture to blood flash was 115 (Ϯ184) seconds for the US group vs. 512 (Ϯ698) seconds for the landmark group (p < 0.0001). The mean number of CVA attempts in the US group was 1.6 (Ϯ1.0) vs. 3.5 (Ϯ2.7) in the landmark group (p < 0.0001). Acute complications were comparable between groups. Comparisons for time, number of CVA attempts, and complications were done specifically for a subset of patients considered to be ''difficult stick'' due to predefined criteria regarding body habitus or vascular disease. Patients considered to be ''difficult sticks'' required a significantly longer amount of time (p < 0.001) for CVA via the landmark technique than patients considered to be ''difficult sticks'' who had CVA with ultrasonic guidance. Time to line placement for the landmark group was 462.7 (Ϯ627) seconds vs. 93.3 (Ϯ176) seconds in the US group. Comparing the same subset also revealed an increase in number of required CVA attempts for the landmark technique group. The number of acute complications in the ''difficult stick'' patients did not show statistical significance (p = 1.00). The landmark group had 60% ''difficult sticks,'' while the ultrasound group had 80%, although the difference was not statistically significant (p = 0.08). Conclusions: Emergency physicians with limited training and experience are able to use ultrasound as an adjunct for central venous access. Ultrasound technology may decrease the number of CVA attempts required to cannulate a central vein and will decrease the amount of time required to cannulate the vein starting from the time when the needle is on the skin, after the ultrasound machine has been set up and turned on. These results are especially true for those patients considered to be ''difficult sticks.''

A signal peptide is required for entry of a preprotein into the secretory pathway, but how it functions in concert with the other transport components is unknown. In Escherichia coli, SecA is a key component of the translocation machinery found in the cytoplasm and at membrane translocation sites. Synthetic signal peptides corresponding to the wild type alkaline phosphatase signal sequence and three sets of model signal sequences varying in hydrophobicity and amino-terminal charge were generated. These were used to establish the requirements for interaction with SecA. Binding to SecA, modulation of SecA conformations sensitive to protease, and stimulation of SecA-lipid ATPase activity occur with functional signal sequences but not with transport-incompetent ones. The extent of SecA interaction is directly related to the hydrophobicity of the signal peptide core region. For signal peptides of moderate hydrophobicity, stimulation of the SecA-lipid ATPase activity is also dependent on amino-terminal charge. The results demonstrate unequivocally that the signal peptide, in the absence of the mature protein, interacts with SecA in aqueous solution and in a lipid bilayer. We show a clear parallel between the hierarchy of signal peptide characteristics that promote interaction with SecA in vitro and the hierarchy of those observed for function in vivo.

Although it is known that virtually all exported proteins require a signal peptide, it is not clearly understood how the signal peptide interfaces with the translocation machinery to achieve transport. In this study we document a direct interaction between the signal peptide and SecA, a primary component of the translocase in Escherichia coli, and show that the signal peptide itself can stimulate SecA-lipid ATPase activity. Using synthetic signal peptides corresponding to the wild type alkaline phosphatase signal sequence and two model sequences, we find that the extent of stimulation