tailieunhanh - báo cáo khoa học: "Molecular understanding of sterically controlled compound release through an engineered channel protein (FhuA)"

Tuyển tập báo cáo các nghiên cứu khoa học quốc tế ngành y học dành cho các bạn tham khảo đề tài: Molecular understanding of sterically controlled compound release through an engineered channel protein (FhuA) | Guven et al. Journal of Nanobiotechnology 2010 8 14 http content 8 1 14 RESEARCH JOURNAL OF NANOBIOTECHNOLOGY Open Access Molecular understanding of sterically controlled compound release through an engineered channel protein FhuA Arcan Guven1 2 Marco Fioroni2 Bernhard Hauer3 and Ulrich Schwaneberg 2 Abstract Background Recently we reported a nanocontainer based reduction triggered release system through an engineered transmembrane channel FhuA A1-160 Onaca etal. 2008 . Compound fluxes within the FhuA A1-160 channel protein are controlled sterically through labeled lysine residues label 3- 2-pyridyldithio propionic-acid-N-hydroxysuccinimide-ester . Quantifying the sterical contribution of each labeled lysine would open up an opportunity for designing compound specific drug release systems. Results In total 12 FhuA A1-160 variants were generated to gain insights on sterically controlled compound fluxes Subset A six FhuA A1-160 variants in which one of the six lysines in the interior of FhuA A1-160 was substituted to alanine and Subset B six FhuA A1-160 variants in which only one lysine inside the barrel was not changed to alanine. Translocation efficiencies were quantified with the colorimetric TMB 3 3 5 5 -tetramethylbenzidine detection system employing horseradish peroxidase HRP . Investigation of the six subset A variants identified position K556A as sterically important. The K556A substitution increases TMB diffusion from 15 to 97 nM s and reaches nearly the TMB diffusion value of the unlabeled FhuA A1-160 102 nM s . The prominent role of position K556 is confirmed by the corresponding subset B variant which contains only the K556 lysine in the interior of the barrel. Pyridyl labeling of K556 reduces TMB translocation to 16 nM s reaching nearly background levels in liposomes 13 nM s . A first B-factor analysis based on MD simulations confirmed that position K556 is the least fluctuating lysine among the six in the channel interior of .