Urease activity in the gastrointestinal tract of the European hare (<Emphasis Type=Italic>Lepus europaeus</Emphasis>)

The intensity of urea recycling in the wild herbivorous European hare has been investigated. The intensity of urease activity in the gastrointestinal tract has been selected as a convenient quantitative measure of nitrogen recycling. High urease activity was detected in the large intestine; it was higher than the previously detected activity in other mammals with postgastric fermentation: pigs, rats, and rabbits. The strong seasonal dynamics of urease activity has been noted: it increases during winter diet consumption, which is poor in available nitrogen.     

Evidence that tartrate-resistant acid phosphatases from osteoclastomas and hairy cell leukemia spleen are members of a multigene family.

1. Osteoclasts and hairy cell leukemia spleen both contain large amounts of a band 5-tartrate-resistant acid phosphatase (TrACP). 2. We have recently purified to homogeneity a band 5 TrACP from human osteoclastomas and two isoforms of band 5 TrACP (5a and 5b) from the spleen of a patient with hairy cell leukemia. 3. Although the N-terminal amino acid sequences and the apparent molecular weights of the osteoclastoma, hairy cell leukemia spleen TrACPs were identical, there were several differences in the physical and biochemical properties between the three isoenzymes. 4. Based on these findings, it is concluded that these isoenzymes are different enzymes, but that they could have originated from a similar ancestral gene. 5. It is proposed that the osteoclastoma and hairy cell leukemia band 5 TrACPs are members of a multigene family.     

A multi-endpoint matched molecular pair (MMP) analysis of 6-membered heterocycles

Aromatic rings, ubiquitous in pharmaceutical compounds, are often exchanged with another ring during the optimization process of drug discovery. Inevitably, the preferred ring system for one endpoint may prove detrimental to another, thus necessitating a holistic, multiple endpoint optimization approach for finding the ideal replacement. Accordingly, we conducted an extensive matched molecular pair (MMP) analysis of common 6-membered aromatic rings across 4 endpoints critical for drug discovery (log D lipophilicity, microsomal metabolism, P-gp efflux and passive permeability). We also investigated the effect of context by considering the connecting atom. Heat maps were created as a simple yet comprehensive way to view and analyze the vast amount of interrelated data. Paired difference statistical tests were used to identify transforms with changes that were significantly different from zero. We conclude that the heat maps of transforms provide a unique and powerful approach for multiparameter optimization.     

The Position of His-Tag in Recombinant OspC and Application of Various Adjuvants Affects the Intensity and Quality of Specific Antibody Response after Immunization of Experimental Mice

Lyme disease, Borrelia burgdorferi-caused infection, if not recognized and appropriately treated by antibiotics, may lead to chronic complications, thus stressing the need for protective vaccine development. The immune protection is mediated by phagocytic cells and by Borrelia-specific complement-activating antibodies, associated with the Th1 immune response. Surface antigen OspC is involved in Borrelia spreading through the host body. Previously we reported that recombinant histidine tagged (His-tag) OspC (rOspC) could be attached onto liposome surfaces by metallochelation. Here we report that levels of OspC-specific antibodies vary substantially depending upon whether rOspC possesses an N'' or C'' terminal His-tag. This is the case in mice immunized: (a) with rOspC proteoliposomes containing adjuvants MPLA or non-pyrogenic MDP analogue MT06; (b) with free rOspC and Montanide PET GEL A; (c) with free rOspC and alum; or (d) with adjuvant-free rOspC. Stronger responses are noted with all N''-terminal His-tag rOspC formulations. OspC-specific Th1-type antibodies predominate post-immunization with rOspC proteoliposomes formulated with MPLA or MT06 adjuvants. Further analyses confirmed that the structural features of soluble N'' and C'' terminal His-tag rOspC and respective rOspC proteoliposomes are similar including their thermal stabilities at physiological temperatures. On the other hand, a change in the position of the rOspC His-tag from N'' to C'' terminal appears to affect substantially the immunogenicity of rOspC arguably due to steric hindrance of OspC epitopes by the C'' terminal His-tag itself and not due to differences in overall conformations induced by changes in the His-tag position in rOspC variants.     

Characterization of Staphylococcus aureus from Humans and a Comparison with ?solates of Animal Origin,in North Dakota,United States

Different clones of methicillin-susceptible (MSSA) and methicillin-resistant (MRSA) Staphylococcus aureus have been found in humans as well as in animals and retail meat. However, more information about the genetic characteristics and similarities between strains is needed. The aim of this study was to identify and characterize Staphylococcus aureus from humans, and to compare their characteristics with isolates of animal origin. A total of 550 nasal swabs were taken from healthy humans, and S. aureus was isolated and identified. Positive S. aureus isolates were subjected to molecular typing and susceptibility testing. In addition, 108 MRSA isolates recovered from clinical patients in the state of North Dakota and 133 S. aureus isolates from animals and meat previously analyzed were included. The nasal carriage of S. aureus in healthy people was 7.6% and, in general, clones were genetically diverse. None of the S. aureus strains obtained from healthy people were mecA- or PVL-positive. A total of 105 (97.2%) MRSA isolates from clinical cases harbored the mecA gene and 11 (10.2%) isolated from blood stream infections harbored the PVL gene. The most common resistance profile among S. aureus from healthy people was penicillin, and from clinical cases were erythromycin-penicillin-ciprofloxacin. The rate of multidrug resistance (MDR) was 70% in humans. Most of the S. aureus harboring mecA and PVL genes were identified as ST5 and ST8, and exhibited MDR. However, S. aureus isolates of animal origin used for comparison exhibited a lower rate of MDR. The most common resistance profiles in isolates of animal origin were penicillin-tetracycline and penicillin-tetracycline-erythromycin, in animals and raw meat, respectively. The ST5 was also found in animals and meat, with ST9 and ST398 being the major clones. The genetic similarity between clones from humans and meat suggests the risk of spread of S. aureus in the food chain.     

Expression of the Na+-d-Glucose Cotransporter SGLT1 in Neurons

Abstract: In brains of the rabbit, pig, and human, expression of the high-affinity Na⁺-d -glucose cotransporter SGLT1 and of the protein RS1, which alters the activity of SGLT1, was demonstrated. In situ hybridization showed that SGLT1 and RS1 are transcribed in pyramidal cells of brain cortex and hippocampus and in Purkinje cells of cerebellum. In neurons of pig brain SGLT1 protein was demonstrated by western blotting with synaptosomal membranes and by immunohistochemistry, which showed SGLT1 in pyramidal and Purkinje cells. To test whether SGLT1 in neurons may be activated during increased d -glucose consumption, an epileptic seizure was induced in rat brain, and the uptake of specific nonmetabolized substrates of SGLT1 {[¹⁴C]methyl-α-d -glucopyranoside ([¹⁴C]AMG)} and of Na⁺-independent transporters {2-deoxy-d -[¹⁴C]glucose([¹⁴C]2-DG)} was analyzed by autoradiography. During the seizure the uptake of AMG and 2-DG was increased in the focus. Within two hours after the seizure 2-DG uptake in the focus returned to normal. In contrast, the AMG uptake in the focus area was still increased 1 day later. The data show that the high-affinity Na⁺-d -glucose cotransporter SGLT1 is expressed in neurons and can be up-regulated.     

Nuclear magnetic resonance-based quantification of organic diphosphates

Phosphorylated compounds are ubiquitous in life. Given their central role, many such substrates and analogs have been prepared for subsequent evaluation. Prior to biological experiments, it is typically necessary to determine the concentration of the target molecule in solution. Here we describe a method where concentrations of stock solutions of organic diphosphates and bisphosphonates are quantified using ³¹P nuclear magnetic resonance (NMR) spectroscopy with standard instrumentation using a capillary tube with a secondary standard. The method is specific and is applicable down to a concentration of 200 μM. The capillary tube provides the reference peak for quantification and deuterated solvent for locking.