Mizrahi V, Usdin M T, Harington A, Dudding L R
Molecular Biology Laboratory, School of Pathology, South African Institute for Medical Research, Johannesburg.
Nucleic Acids Res. 1990 Sep 25;18(18):5359-63. doi: 10.1093/nar/18.18.5359.
Substitution of the conserved Asp-443 residue of HIV-1 reverse transcriptase by asparagine specifically suppressed the ribonuclease H activity of the enzyme without affecting the reverse transcriptase activity, suggesting involvement of this ionizable residue at the ribonuclease H active site. An analogous asparagine substitution of the Asp-498 residue yielded an unstable enzyme that was difficult to enzymatically characterize. However, the instability caused by the Asn-498 mutation was relieved by the introduction of a second distal Asn-443 substitution, yielding an enzyme with wild type reverse transcriptase activity, but lacking ribonuclease H activity.
将HIV-1逆转录酶保守的天冬氨酸-443残基替换为天冬酰胺，特异性地抑制了该酶的核糖核酸酶H活性，而不影响逆转录酶活性，这表明这个可电离残基参与了核糖核酸酶H活性位点的构成。将天冬氨酸-498残基进行类似的天冬酰胺替换，得到一种不稳定的酶，难以对其进行酶学特性鉴定。然而，天冬酰胺-498突变引起的不稳定性通过引入第二个远端天冬酰胺-443替换得以缓解，产生了一种具有野生型逆转录酶活性但缺乏核糖核酸酶H活性的酶。
