High-level overproduction of His-tagged Tth DNA polymerase in Thermus thermophilus

Article Abstract:

A new plasmid is described, which uses a modified version of the promoter from the respiratory nitrate reductase (narp) to overproduce active DNA polymerase from Thermus thermophilus in larger amounts than those obtained with T7-dependent expression systems of Escherichia coli. It is concluded that this new expression system represents a competitive route for the direct overexpression of soluble enzymes in T. thermophilus, even compared to the high-level overexpression system of E. coli.

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Titration and conditional knockdown of the prfB gene in Escherichia ccoli: Effects on growth and overproduction of the recombinant mammalian selenoprotein thioredoxin reductase

Article Abstract:

An Escherichia coli strain is constructed that has endogenous chromosomal promoter of prfB replaced with the titratable [P.sub.BAD] promoter. The analysis has shown that expression of prfB can be titrated through targeted exchange of the native promoter with a [P.sub.BAD]-promoter and that knockdown of release factor 2 (RF2) has resulted in almost full efficiency of selenocysteine (Sec) incorporation at the cost of lower total seleno-protein yield.

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Use of a dominant rpsL allele conferring streptomycin dependence for positive and negative selection in thermos thermophilus

Article Abstract:

A spontaneous rpsL mutant of Thermus thermophilus is isolated to search a new selection marker for this organism. Models built on the available three-dimensional structures of the 30S ribosomal subunit revealed that the K47R mutation directly affects the streptomycin binding site on S12.

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