Association of the components of the binary toxin from Bacillus sphaericus in solution and with model lipid bilayers.

We show herein that interaction in aqueous solution of the two components of binary toxin from Bacillus sphaericus, BinA and BinB, leads to a dramatic conformational change, from beta turns or random coil, to beta structure. Also, either BinA or BinB separately or their equimolar mixture, interact with lipid bilayers resulting in further conformational changes. Upon membrane association, the change in conformation observed for BinA or BinB separately is different from that observed when the proteins are combined, indicating that proper folding depends on the presence of the complementary subunit. We also show, in contrast to previous reports, that BinB, but not BinA, is able to insert in model neutral lipid monolayers.

Engagement of Na,K-ATPase beta3 subunit by a specific mAb suppresses T and B lymphocyte activation.

In order to identify new molecules involved in regulation of T cell proliferation, we generated various mAb by immunization of mice with the T cell line Molt4. We found one mAb (termed P-3E10) that down-regulated the in vitro T cell proliferation induced by CD3-specific OKT3 mAb. The P-3E10 mAb was also able to inhibit IFN-gamma, IL-2, IL-4 and IL-10 production of OKT3-activated T cells. The antigen recognized by P-3E10 mAb is broadly expressed on all hematopoietic as well as on all non-hematopoietic cell lines tested so far. Within peripheral blood leukocytes, the P-3E10 antigen was detected on lymphocytes, monocytes and granulocytes. Human umbilical vein endothelial cells (HUVEC) also scored positively. By evaluating the effect of P-3E10 mAb on these cell types we found that it also inhibited anti-IgM-induced B cell proliferation. However, it did not block growth factor-mediated proliferation of HUVEC, and spontaneous proliferation of SupT-1, Jurkat, Molt4 and U937 cell lines. Moreover, it did not influence phagocytosis of human blood monocytes and granulocytes. Biochemical analysis revealed that the P-3E10 antigen is a protein with a mol. wt of 45-50 kDa under non-reducing and 50-55 kDa under reducing conditions. By using a retroviral cloning system, the P-3E10 antigen was cloned. Sequence analysis revealed the P-3E10 antigen to be identical to the beta3 subunit of the Na,K-ATPase.