Bicine

Fibrillar aggregates of amyloid-β (Aβ) are the main component of plaques lining the cerebrovasculature in cerebral amyloid angiopathy. As the predominant Aβ isoform in vascular deposits, Aβ40 is a valuable target in cerebral amyloid angiopathy research. However, the slow process of Aβ40 aggregation in vitro is a bottleneck in the search for Aβ-targeting molecules. In this study, we sought a method to accelerate the aggregation of Aβ40 in vitro, to improve experimental screening procedures. We evaluated the aggregating ability of bicine, a biological buffer, using various in vitro methods. Our data suggest that bicine promotes the aggregation of Aβ40 with high speed and reproducibility, yielding a mixture of aggregates with significant β-sheet-rich fibril formation and toxicity.

The toxicity of free Mn(2+) is a bottleneck for the in vivo application of manganese ion enhanced MRI. To reduce free Mn(2+) concentration ([Mn(2+) ]), a low affinity chelate reagent: N,N-bis(2-hydroxyethyl)glycine (bicine) was used. Considering the conditional association constant of Mn-bicine at pH 7.4 (10(2.9) M(-1) ), (i) a 100 mM Mn-bicine solution should contain about 10 mM of free manganese ion, but (ii) free manganese will make up 3/4 of the final plasma concentration (0.5 mM) with an intravenous infusion of 100 mM Mn-bicine. The T(1) relaxivity of Mn-bicine in a 5 mM Mn-bicine solution was estimated as 5 mM(-1) sec(-1) at 24°C, 7 T in a pH range of 6.8-7.5. Mn-bicine demonstrated a tendency for better contractility when employed with an isolated perfused frog heart, compared with MnCl(2) . A venous infusion of 100 mM Mn-bicine (8.3 μmol kg(-1) min(-1) ) showed a minimal decrease and maintained a constant heart rate level and arterial pressure in rats, while rats infused with 100 mM of MnCl(2) showed a significant suppression of the hemodynamic functions. Thus, Mn-bicine appears to be a better choice for maintaining the vital conditions of experimental animals, and may improve the reproducibility of manganese ion enhanced MRI.

A novel, Bicine-based SDS-PAGE buffer system was developed for the analysis of membrane proteins. The method involves molecular weight-based separations of fully denatured and solubilized proteins in two dimensions. This doubled SDS-PAGE (dSDS-PAGE) approach produced a diagonal arrangement of protein spots and successfully circumvented problems associated with membrane proteome analysis involving traditional gel-based methods. Membrane proteins from the anaerobic bacterium Clostridium thermocellum were used for these investigations. Tricine-dSDS-PAGE and the newly developed Bicine-dSDS-PAGE were compared with the standard glycine-dSDS-PAGE (Laemmli protocol) in their suitability to separate C. thermocellum membrane proteins. Large-format gel experiments using optimized gel preparation and running buffer conditions revealed a 112% increase in protein spot count for Tricine-dSDS-PAGE and a 151% increase for Bicine-dSDS-PAGE, compared to glycine-dSDS-PAGE. The data clearly indicated that Bicine-dSDS-PAGE is a superior method for the analysis of membrane proteins, providing enhanced resolution and protein representation.