Streptavidin maleimide-activated

Immediately purify the maleimide-activated avidin or streptavidin away from excess cross-linker and reaction by-products by gel filtration on a desalting column (Sephadex G-25 or the equivalent). Use 0.1 M sodium phosphate, 0.15 M NaCl, pH 7.2, as the chromatography buffer. Pool the fractions containing protein (the first peak eluting from the column). After elution, adjust the protein concentration to 10 mg/ml for the conjugation reaction (centrifugal concentrators work well for this step). At this point, the maleimide-activated avidin may be frozen and lyophilized to preserve its maleimide activity. The modified pro-... 

Immediately mix the thiolated enzyme with an amount of maleimide-activated avidin (or streptavidin) to obtain the desired molar ratio of enzyme to avidin in the conjugate. Use of a 4 1 (enzyme avidin) molar ratio in the conjugation... 

A biotin-linked maleimide derivative was used as the reactive partner in the Diels-Alder reaction. These compounds were reacted in the presence of appropriate metal ions. The products of the Diels-Alder reaction contained both biotin and an RNA chain. The products with biotin attached were selected through specific binding with streptavidin. The RNA sequences of interest show catalytic activity towards the Diels-Alder reaction. These were transcribed into DNA form. The DNA sequences obtained were multiphed by PGR treatment, and a new RNA library was synthesized from these DNA sequences. Repeating the reaction / selection process and PGR treatment yielded RNA sequences that were optimized for catalyzing the Diels-Alder reaction. 

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