Substitution of a highly basic helix/loop sequence into the RNase H domain of human immunodeficiency virus reverse transcriptase restores its Mn(2+)-dependent RNase H activity.


Abstract

Human immunodeficiency virus (HIV) reverse transcriptase (RT) is a multifunctional protein, containing both DNA polymerase and RNase H activity. The RNase H activity of HIV RT catalyzes the hydrolysis of the RNA strand of RNA.DNA hybrids. While the domain that carries out the RNase H activity in HIV RT can be expressed as an independent, folded polypeptide, it is inactive as an RNase H. Here, we report the overexpression and purification of an active, recombinant HIV RNase H domain in which the sequence corresponding to the Escherichia coli RNase H1 basic helix/loop has been substituted for the corresponding sequence of HIV RNase H. The resulting polypeptide (RNH102) has Mn(2+)-dependent RNase H activity and is more stable than the independently expressed wild-type HIV RNase H domain. Study holds ProTherm entries: 2760 Extra Details: ribonuclease HI; Human immunodeficiency virus; HIV;,Escherichia coli; activity

Submission Details

ID: B5vH4TCA

Submitter: Connie Wang

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

Version: 1

Publication Details
Keck JL;Marqusee S,Proc. Natl. Acad. Sci. U.S.A. (1995) Substitution of a highly basic helix/loop sequence into the RNase H domain of human immunodeficiency virus reverse transcriptase restores its Mn(2+)-dependent RNase H activity. PMID:7535929
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
4Z0U 2015-03-26T00:00:00+0000 2.0 RNase HI/SSB-Ct complex
1F21 2000-05-22T00:00:00+0000 1.4 DIVALENT METAL COFACTOR BINDING IN THE KINETIC FOLDING TRAJECTORY OF E. COLI RIBONUCLEASE HI
1G15 2000-10-10T00:00:00+0000 1.9 CO-CRYSTAL OF E. COLI RNASE HI WITH TWO MN2+ IONS BOUND IN THE THE ACTIVE SITE
1GOA 1993-05-10T00:00:00+0000 1.9 COOPERATIVE STABILIZATION OF ESCHERICHIA COLI RIBONUCLEASE HI BY INSERTION OF GLY-80B AND GLY-77-> ALA SUBSTITUTION
1GOB 1993-05-10T00:00:00+0000 2.0 COOPERATIVE STABILIZATION OF ESCHERICHIA COLI RIBONUCLEASE HI BY INSERTION OF GLY-80B AND GLY-77-> ALA SUBSTITUTION
1GOC 1993-05-10T00:00:00+0000 2.0 COOPERATIVE STABILIZATION OF ESCHERICHIA COLI RIBONUCLEASE HI BY INSERTION OF GLY-80B AND GLY-77-> ALA SUBSTITUTION
1JL1 2001-07-13T00:00:00+0000 1.3 D10A E. coli ribonuclease HI
1JL2 2001-07-13T00:00:00+0000 1.76 Crystal structure of TCEO RNase H-a chimera combining the folding core from T. thermophilus RNase H and the remaining region of E. coli RNase H
1JXB 2001-09-06T00:00:00+0000 1.6 I53A, a point mutant of the cysteine-free variant of E. coli Rnase HI
1KVA 1996-10-04T00:00:00+0000 1.8 E. COLI RIBONUCLEASE HI D134A MUTANT

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
90.9 Ribonuclease HI A6T512 RNH_KLEP7
90.9 Ribonuclease HI B5Y1G2 RNH_KLEP3
92.9 Ribonuclease HI C0Q6N2 RNH_SALPC
93.5 Ribonuclease HI P0A2B9 RNH_SALTY
93.5 Ribonuclease HI P0A2C0 RNH_SALTI
93.5 Ribonuclease HI B4TYH0 RNH_SALSV
93.5 Ribonuclease HI B5BDW5 RNH_SALPK
93.5 Ribonuclease HI A9MZ19 RNH_SALPB
93.5 Ribonuclease HI Q5PFD8 RNH_SALPA
93.5 Ribonuclease HI B4SV39 RNH_SALNS
93.5 Ribonuclease HI B4TK85 RNH_SALHS
93.5 Ribonuclease HI B5R5L3 RNH_SALG2
93.5 Ribonuclease HI B5R449 RNH_SALEP
93.5 Ribonuclease HI B5FJ58 RNH_SALDC
93.5 Ribonuclease HI Q57SZ6 RNH_SALCH
93.5 Ribonuclease HI A9MPF1 RNH_SALAR
93.5 Ribonuclease HI B5F8X2 RNH_SALA4
93.5 Ribonuclease HI A8AKR0 RNH_CITK8
99.4 Ribonuclease HI Q0TLC3 RNH_ECOL5
99.4 Ribonuclease HI A7ZWF6 RNH_ECOHS
99.4 Ribonuclease HI B7LHC0 RNH_ECO55
99.4 Ribonuclease HI B7UJB0 RNH_ECO27
100.0 Ribonuclease HI Q3Z5E9 RNH_SHISS
100.0 Ribonuclease HI P0A7Y7 RNH_SHIFL
100.0 Ribonuclease HI Q32JP9 RNH_SHIDS
100.0 Ribonuclease HI Q325T2 RNH_SHIBS
100.0 Ribonuclease HI B2U352 RNH_SHIB3
100.0 Ribonuclease HI B7LW89 RNH_ESCF3
100.0 Ribonuclease HI B1LHM3 RNH_ECOSM
100.0 Ribonuclease HI B6HZS7 RNH_ECOSE
100.0 Ribonuclease HI B7N876 RNH_ECOLU
100.0 Ribonuclease HI P0A7Y4 RNH_ECOLI
100.0 Ribonuclease HI B1IPU4 RNH_ECOLC
100.0 Ribonuclease HI P0A7Y5 RNH_ECOL6
100.0 Ribonuclease HI B1XD78 RNH_ECODH
100.0 Ribonuclease HI C4ZRV1 RNH_ECOBW
100.0 Ribonuclease HI B7M213 RNH_ECO8A
100.0 Ribonuclease HI B7MQ23 RNH_ECO81
100.0 Ribonuclease HI B7NKW4 RNH_ECO7I
100.0 Ribonuclease HI B5Z0I8 RNH_ECO5E
100.0 Ribonuclease HI P0A7Y6 RNH_ECO57
100.0 Ribonuclease HI B7MBJ0 RNH_ECO45
100.0 Ribonuclease HI A7ZHV1 RNH_ECO24