Table 2.

Deacylase activity in crude extracts

Strain	Deacylase activity (U/mg)
K37ΔrecAλDE3 + pET3alpa	0.9
K37ΔrecAΔtyrHλDE3 + pET3alpa	<0.1
K37ΔrecAΔtyrHλDE3 + pET3alpa::GEK1	7700
K37ΔrecAλDE3 + pET15blpa	1.0
K37ΔrecAΔtyrHλDE3 + pET15blpa	<0.1
K37ΔrecAΔtyrHλDE3 + pET15blpa::GEK1	4000

Cells were grown overnight at 37°C in 2xTY medium with 100 μg/ml ampicillin. Then, IPTG was added at a final concentration of 1 mM, and growth was continued for 5 h at room temperature. Specific deacylase activities were measured in crude extracts obtained by sonication. Final total protein concentration in the extracts was 10–20 mg/ml. Rates of hydrolysis of d-[³H]Tyr-tRNA^Tyr in the presence of 20 mM Tris-HCl (pH 7.8), 100 nM d-[³H]Tyr-tRNA^Tyr, 5 mM MgCl₂, 40 μM zinc acetate, 50 μg/ml BSA and 2.5 mM 2-mercaptoethanol were measured as described in Materials and Methods. One unit corresponds to the enzyme activity capable of hydrolyzing 1 pmol of d-[³H]Tyr-tRNA^Tyr per min under the above assay conditions.