Functional interactions between putative intramembrane charged residues in the lactose permease of Escherichia coli.

Using a lactose permease mutant devoid of Cys residue ("C-less permease"), we systematically replaced putative intramembrane charged residues with Cys. Individual replacements for Asp-237, Asp-240, Glu-269, Arg-302, Lys-319, His-322, Glu-325, or Lys-358 abolish active lactose transport. When Asp-237 and Lys-358 are simultaneously replaced with Cys and/or Ala, however, high activity is observed. Therefore, when either Asp-237 or Lys-358 is replaced with a neutral residue, leaving an unpaired charge, the permease is inactivated, but neutral replacement of both residues yields active permease [King, S. C., Hansen, C. L. & Wilson, T. H. (1991) Biochim. Biophys. Acta 1062, 177-186]. Remarkably, moreover, when Asp-237 is interchanged with Lys-358, high activity is observed. The observations provide a strong indication that Asp-237 and Lys-358 interact to form a salt bridge. In addition, the data demonstrate that neither residue nor the salt bridge plays an important role in the transport mechanism. Thirteen additional double mutants were constructed in which a negative and a positively charged residue were replaced with Cys. Only Asp-240-->Cys/Lys-319-->Cys exhibits significant activity, accumulating lactose to 25-30% of the steady state observed with C-less permease. Replacing either Asp-240 or Lys-319 individually with Ala also inactivates the permease, but double mutants with neutral substitutions (Cys and/or Ala) at both positions exhibit essentially the same activity as Asp-240-->Cys/Lys-319-->Cys. In marked contrast to Asp-237 and Lys-358, interchanging Asp-240 and Lys-319 abolishes active lactose transport. The results demonstrate that Asp-240 and Lys-319, like Asp-237 and Lys-358, interact functionally and may form a salt bridge. However, the interaction between Asp-240 and Lys-319 is clearly more complex than the interaction between Asp-237 and Lys-358. In any event, the findings suggest that putative transmembrane helix VII lies next to helices X and XI in the tertiary structure of lactose permease.     

Spontaneous loss of permanent teeth: a case report

Pathological root resorption has clinical symptoms of great importance when it appears in permanent teeth, as its etiology is not always clear. It is important to examine all local and general factors before determining its "idiopathic" cause. A case of spontaneous loss of permanent teeth in a 19 year-old female is presented. Through a meticulous study correlating the clinical history and the evolution of the symptoms, we can suspect that non-dental medical interventions requiring general anesthesia can lead to the loss of permanent teeth through root resorption.     

Linkage of the MHC to familial multiple sclerosis suggests genetic heterogeneity. The Multiple Sclerosis Genetics Group

Multiple sclerosis (MS) is a demyelinating autoimmune disease of the central nervous system. While its etiology is not well understood, genetic factors are clearly involved. Until recently, most genetic studies in MS have been association studies using the case-control design testing specific candidate genes and studying only sporadic cases. The only consistently replicated finding has been an association with the HLA-DR2 allele within the major histocompatibility complex (MHC) on chromosome 6. Using the genetic linkage design, however, evidence for and against linkage of the MHC to MS has been found, fostering suggestions that sporadic and familial MS have different etiologies. Most recently, two of four genomic screens demonstrated linkage to the MHC, although specific allelic associations were not tested. Here, a dataset of 98 multiplex families was studied to test for an association to the HLA-DR2 allele in familial MS and to determine if genetic linkage to the MHC was due solely to such an association. Three highly polymorphic markers (HLA-DR, D6S273 and TNFbeta) in the MHC demonstrated strong genetic linkage (parametric lod scores of 4.60, 2.20 and 1.24, respectively) and a specific association with the HLA-DR2 allele was confirmed (TDT; P < 0.001). Stratifying the results by HLA-DR2 status showed that the linkage results were limited to families segregating HLA-DR2 alleles. These results demonstrate that genetic linkage to the MHC can be explained by the HLA-DR2 allelic association. They also indicate that sporadic and familial MS share a common genetic susceptibility. In addition, preliminary calculations suggest that the MHC explains between 17 and 62% of the genetic etiology of MS. This heterogeneity is also supported by the minority of families showing no linkage or association with loci within the MHC.      

Granulocyte turnover in the feline intestine

The objective of this study was to determine the turnover rate of the extravascular pool of granulocytes in different regions of the feline gastrointestinal tract. Leukocyte emigration from the vasculature was prevented over a 48-h period by repeated intravenous injections of a monoclonal antibody (MAb IB₄) directed against the leukocyte adhesion glycoprotein complex CD11/CD18. Tissue-associated myeloperoxidase (MPO) activity was used to monitor the total tissue granulocyte pool at 0.5, 12, 24, and 48 h after MAb IB₄ administration. The mucosal layer of the duodenum, jejunum, ileum, and colon exhibited different kinetics of granulocyte clearance, with average life-spans (t1/2) ranging between 6.9 (colon) and 10.4 h (duodenum). Granulocyte clearance rates of 0.5 × 10⁶ and 2.4 x 10⁶ cells/h/g tissue were estimated (from measured values oft1/2 and tissue granulocyte pool) for the small bowel and colonie mucosae, respectively. The submucosal layer of the intestine exhibited a biphasic reduction in tissue MPO activity following immunoneutralization of CD11/CD18, with an initialt1/2 0.5 h followed by at1/2 of 36–60 h. The initial rapid decline in tissue MPO suggests that a significant fraction of granulocytes in the submucosa is localized in a readily exchangeable pool (e.g., marginated cells within the vasculature). The results of this study indicate that the average life-span of resident granulocytes varies significantly between different regions of the gastrointestinal tract, with the intestinal mucosa exhibiting at1/2 comparable to that previously reported for circulating feline neutrophils (R 8 h).     

Salmonella-specific monoclonal antibodies against recombinant Salmonella typhi 36-kilodalton porin.

Mouse monoclonal antibodies were raised against recombinant Salmonella typhi 36-kDa porin monomer. Specificities of 16 monoclonal antibodies were analyzed as reactivity patterns in dot immunobinding and Western blot (immunoblot) assays using isolated outer membrane proteins of gram-negative bacteria and cloned purified S. typhi porin monomers and trimers. Four monoclonal antibodies were specific for Salmonella spp.