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. 1990 Sep 17;270(1-2):76-80.
doi: 10.1016/0014-5793(90)81238-j.

Purification and Characterization of the RNase H Domain of HIV-1 Reverse Transcriptase Expressed in Recombinant Escherichia Coli

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Purification and Characterization of the RNase H Domain of HIV-1 Reverse Transcriptase Expressed in Recombinant Escherichia Coli

S P Becerra et al. FEBS Lett. .
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Abstract

The ribonuclease H (RNase H) domain of human immuno-deficiency virus (HIV-1) reverse transcriptase has been produced with the aim of providing sufficient amounts of protein for biophysical studies. A plasmid vector is described which directs high level expression of the RNase H domain under the control of the lambda PL promoter. The domain corresponds to residues 427-560 of the 66 kDa reverse transcriptase. The protein was expressed in Escherichia coli and was purified using ion-exchange and size exclusion chromatography. The purified protein appears to be in a native-like homogeneous conformational state as determined by 1H-NMR spectroscopy and circular dichroism measurements. HIV-protease treatment of the RNase H domain resulted in cleavage between Phe-440 and Tyr-441.

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