کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1952751 | 1057229 | 2010 | 7 صفحه PDF | دانلود رایگان |
A magnetic sensor technique was applied to analyze the interaction of immobilized bacterial RNase P protein and 3′-biotinylated RNase P RNA bound to streptavidin-coated magnetic beads. Our measurements with three types of beads from different suppliers resulted in Kd values of about 1–2 nM (at 4.5 mM Mg2+ and 150 mM NH4+) for Escherichia coli RNase P RNA and protein, consistent with previous analyses using different techniques. We further measured affinity of the E. coli RNase P protein to chimeric RNase P RNA variants, consisting of an E. coli specificity domain and an engineered archaeal catalytic domain. A “bacterial-like” 1-bp insertion and 2-nt deletion in the helix P2/P3 region largely improved affinity, providing independent evidence that these elements are crucial for interaction of the two RNase P subunits. Moreover, our study documents that the properties of the streptavidin-coated magnetic beads decide on success or failure of the technique.
Journal: Biochimie - Volume 92, Issue 7, July 2010, Pages 772–778