PURIFICATION AND CHARACTERIZATION OF THE RNASE H DOMAIN OF HIV-1 REVERSE-TRANSCRIPTASE EXPRESSED IN RECOMBINANT ESCHERICHIA-COLI

被引：54
作者：
BECERRA, SP
CLORE, GM
GRONENBORN, AM
KARLSTROM, AR
STAHL, SJ
WILSON, SH
WINGFIELD, PT
机构：
[1] NIH,PROT EXPRESS LAB,BETHESDA,MD 20892
[2] NIDDK,CHEM PHYS LAB,BETHESDA,MD 20892
[3] NIH,BIOCHEM LAB,BETHESDA,MD 20892
[4] NHLB,BIOCHEM LAB,BETHESDA,MD 20892
来源：
FEBS LETTERS | 1990年 / 270卷 / 1-2期
关键词：
HIV protease; HIV-1 reverse transcriptase; HIV-1 RNase H; Protein conformation; Protein expression; Protein purification;
D O I：
10.1016/0014-5793(90)81238-J
中图分类号：
Q5 [生物化学]; Q7 [分子生物学];
学科分类号：
071010 ; 081704 ;
摘要：
The ribonuclease H (RNase H) domain of human immune-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 λ 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. © 1990.
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页码：76 / 80
页数：5
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