• DnaK from Bifidobacterium animalis subsp. lactis is a surface-exposed human plasminogen receptor upregulated in response to bile salts.

      Candela, Marco; Centanni, Manuela; Fiori, Jessica; Biagi, Elena; Turroni, Silvia; Orrico, Catia; Bergmann, Simone; Hammerschmidt, Sven; Brigidi, Patrizia; Department of Pharmaceutical Sciences, University of Bologna, Italy. (2010-06)
      Bifidobacterium animalis subsp. lactis lives in the gastrointestinal tract of most mammals, including humans. Recently, for the probiotic strain B. animalis subsp. lactis BI07, a dose-dependent plasminogen-binding activity was demonstrated and five putative plasminogen-binding proteins were identified. Here we investigated the role of surface DnaK as a B. animalis subsp. lactis BI07 plasminogen receptor. DnaK was visualized on the bacterial cell surface by transmission electron microscopy. The His-tagged recombinant DnaK protein showed a high affinity for human plasminogen, with an equilibrium dissociation constant in the nanomolar range. The capability to tolerate physiological concentrations of bile salts is a crucial feature for an intestinal symbiont micro-organism. By proteome analysis we demonstrated that the long-term exposure of B. animalis subsp. lactis BI07 to bile salts results in the upregulation of important surface plasminogen receptors such as DnaK and enolase. Moreover, adaptation of B. animalis subsp. lactis BI07 to physiological concentrations of bile salts significantly increased its capacity to interact with the host plasminogen system. By enhancing the bacterial capacity to interact with the host plasminogen, the gut bile environment may facilitate the colonization of the human host by B. animalis subsp. lactis BI07.
    • Exocytotic process as a novel model for mineralization by osteoblasts in vitro and in vivo determined by electron microscopic analysis.

      Rohde, Manfred; Mayer, H; Department of Microbial Pathogenesis, Helmholtz Center for Infection Research, Inhoffenstrasse 7, D-38124 Braunschweig, Germany. manfred.rohde@helmholtz-hzi.de (2007-05)
      The process of biomineralization has been examined during osteoblastic differentiation of bone marrow stroma cells (BMSCs) from embryonic chick in culture and in periosteum itself by a number of different techniques including transmission and scanning electron microscopy. In cell culture of BMSCs at days 20-25, crystals were accumulated extracellularly in the collagen matrix, resulting in large plate-like crystallites and noncollagen associated on the culture disk surface. In contrast, up to days 10-18, mainly intracellular mineralization was visible by numerous needle-like crystal structures in the cell cytoplasm and in vacuoles. After 20-30 days, the crystal content of these vacuoles is released, most probably by membrane fusion to the outside of the cells. Energy-dispersive X-ray analysis (EDX), electron spectroscopic imaging, and electron energy loss spectroscopy demonstrated that Ca, O, and P are located in the intra- and extracellular needle-like crystals. From EDX spectra a Ca/P ratio of 1.3 was estimated for the intracellular structures and a Ca/P ratio of 1.5, for the extracellular material (for comparison, the Ca/P ratio in tibiae is 1.6). X-ray diffraction and quantitative infrared spectral analyses also demonstrated an increase of crystalline bone apatite along the mineralization process. In addition to the finding in vitro, the presence of intracellular needle-like crystals in vacuoles could be demonstrated in vivo in osteoblastic cells of the periosteum in tibia of day 11. The results are in favor of a novel model for mineralization by osteoblasts, in which amorphous Ca/P material is directly secreted via an exocytotic process from vacuoles of the osteoblast, deposited extracellularly, propagated into the collagen fibril matrix, and matured to hydroxyapatite.
    • On the origin of the electrostatic surface potential of Aspergillus niger spores in acidic environments.

      Wargenau, Andreas; Fleissner, André; Bolten, Christoph Josef; Rohde, Manfred; Kampen, Ingo; Kwade, Arno; Institut für Partikeltechnik, Technische Universität Braunschweig, Volkmaroder Straße 5, D-38104 Braunschweig, Germany. wargenau@a-wargenau.de (2011-12)
      The electrostatic surface potential of fungal spores is generally regarded as potentially influencing spore aggregation and pellet formation in submerged cultures of filamentous fungi. Spores of Aspergillus niger are typically characterized by negative zeta potentials over a wide range of pH values. In this study, this particular behavior is ascribed to the presence of an extensive melanin coating. It is proposed on the basis of zeta potential and pigment extraction experiments that this outermost layer affects the pH-dependent surface potential in two manners: (i) by the addition of negative charges to the spore surface and (ii) by the pH-dependent release of melanin pigment. Chemical analyses revealed that deprotonation of melanin-bound carboxyl groups is most probably responsible for pigment release under acidic conditions. These findings were incorporated into a simple model which has the ability to qualitatively explain the results of zeta potential experiments and, moreover, to provide the basis for quantitative investigations on the role of electrostatics in spore aggregation.