Abstract

The ribonuclease H (RNase H) family of enzymes selectively degrades the RNA strand of RNA-DNA hybrids. This activity is essential for retroviruses such as HIV and resides in a domain of the larger reverse transcriptase molecule. RNase H from Escherichia coli is the best-characterized member of the family and serves as a model for structure/function studies. Despite having almost identical alpha + beta folds, the isolated domain from HIV is inactive and much less stable than the E. coli homolog. The HIV domain also shows increased disorder in its C-terminal regions (E-helix and His-containing loop). We investigated the importance of this region by studying a variant of E. coli RNase H lacking these elements (RNHdeltaE). Despite the elimination of 33 of 155 residues (including a complete helix), this C-terminal deletion mutant folds cooperatively as a subdomain. Surprisingly, this protein lacking residues near the active site retains weak Mn2+-dependent activity. A peptide corresponding to the deleted E-helix is helical in isolation and stimulates the activity of the deletion mutant in vitro. These results have implications for the catalytic mechanism of RNase H and drug design targeted to HIV reverse transcriptase. Study holds ProTherm entries: 2761, 2762 Extra Details: measurement was made in the presence of 0.8M GdnHCl ribonuclease HI; stability; helix; subdomain; HIV;,Escherichia coli; activity

Submission Details

ID: gVxPLfCT

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:19 p.m.

Version: 1

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