Superstoichiometric Binding of the Anticancer Agent Titanocene Dichloride by Human Serum Transferrin and the Accompanying Lobe Closure.

Titanocene dichloride (TDC) is an anticancer agent that delivers Ti(IV) into each of the two Fe(III) binding sites of bilobal human serum transferrin (Tf). This protein has been implicated in the selective transport of Ti(IV) to cells. How Ti(IV) might be released from the Tf Fe(III) binding site has remained a question, and crystal structures have raised issues about lobe occupancy and lobe closure in Ti(IV)-loaded Tf, compared with the Fe(III)-loaded form. Here, inductively coupled plasma optical emission spectroscopy reveals that Tf can stabilize toward hydrolytic precipitation more than 2 equiv of Ti, implying superstoichiometric binding beyond the two Fe(III) binding sites. Further studies support the inability of TDC to induce a complete lobe closure of Tf. Fluorescence data for TDC binding at low equivalents of TDC support an initial protein conformational change and lobe closure upon Ti binding, whereas data at higher equivalents support an open lobe configuration. Spectroscopic titration reveals less intense protein-metal electronic transitions as TDC equivalents are increased. Denaturing urea-PAGE gels and small angle X-ray scattering studies support an open lobe conformation. The concentrations of bicarbonate used in some earlier studies are demonstrated here to cause a pH change over time, which may contribute to variation in the apparent molar absorptivity associated with Ti(IV) binding in the Fe binding site. Finally, Fe(III)-bound holo-Tf still stabilizes TDC toward hydrolytic precipitation, a finding that underscores the importance of the interactions of Tf and TDC outside the Fe(III) binding site and suggests possible new pathways of Ti introduction to cells.

TiO2 exposure alters transition metal ion quota in Rhodococcus ruber GIN-1.

After exposure to micron-sized TiO2 particles, anatase and/or rutile, Rhodococcus ruber GIN-1 accumulates an increased concentration (2.2 ± 0.2 mg kg-1) of mobilized Ti into its biomass with concomitant decreases in cellular biometals Fe, Zn, and possibly Mn, while levels of Cu and Al are unaffected.

Solution structure, glycan specificity and of phenol oxidase inhibitory activity of Anopheles C-type lectins CTL4 and CTLMA2.

Malaria, the world's most devastating parasitic disease, is transmitted between humans by mosquitoes of the Anopheles genus. An. gambiae is the principal malaria vector in Sub-Saharan Africa. The C-type lectins CTL4 and CTLMA2 cooperatively influence Plasmodium infection in the malaria vector Anopheles. Here we report the purification and biochemical characterization of CTL4 and CTLMA2 from An. gambiae and An. albimanus. CTL4 and CTLMA2 are known to form a disulfide-bridged heterodimer via an N-terminal tri-cysteine CXCXC motif. We demonstrate in vitro that CTL4 and CTLMA2 intermolecular disulfide formation is promiscuous within this motif. Furthermore, CTL4 and CTLMA2 form higher oligomeric states at physiological pH. Both lectins bind specific sugars, including glycosaminoglycan motifs with ß1-3/ß1-4 linkages between glucose, galactose and their respective hexosamines. Small-angle x-ray scattering data supports a compact heterodimer between the CTL domains. Recombinant CTL4/CTLMA2 is found to function in vivo, reversing the enhancement of phenol oxidase activity in dsCTL4-treated mosquitoes. We propose these molecular features underline a common function for CTL4/CTLMA2 in mosquitoes, with species and strain-specific variation in degrees of activity in response to Plasmodium infection.