title: Effect of Escherichia coli chaperonin GroELS on heterologously expressed human immunodeficiency virus type 1 reverse transcriptase in vivo and in vitro.
abstract: The two subunits of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (HIV-1 RT), p66 and p51, were coexpressed in Escherichia coli along with the E. coli chaperonin system GroEL/GroES. Coexpression increases the yield of heterodimeric HIV-1 RT by a factor of 4 to 5 and improves the nucleic acid binding affinity of HIV-1 RT by a factor of 1.6. We have analyzed the reasons for the improvements. The total increase in yield of HIV-1 RT can be attributed to an accumulation of RT subunits in the cells (factor of about 2.8) and an increased growth of the E. coli cells (factor of about 1.4). One reason for the accumulation in the cells is an improved stability of HIV-1 RT subunits toward bacterial proteases. In vitro studies showed that the nucleic acid binding affinity of HIV-1 RT purified from cells that did not coexpress GroELS was stimulated by adding purified GroELS (approx 1.5-fold), whereas HIV-1 RT stemming from cells coexpressing GroELS was stimulated only marginally (approx 1.1-fold). The in vivo as well as the in vitro studies suggest that the chaperonin interacts with HIV-1 RT and therefore affects the folding process of HIV-1 RT.
authors.affiliation: Max-Planck-Institut für Biochemie, Martinsried, Germany.
authors.forename: G
authors.initials: G
authors.lastname: Maier
authors.forename: E
authors.initials: E
authors.lastname: Manakova
authors.forename: H
authors.initials: H
authors.lastname: Heumann
mesh_terms: D002455:Cell Division; D018834:Chaperonin 60 / Q000378:metabolism*; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D054303:HIV Reverse Transcriptase / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013997:Time Factors
chemical_list: D018834:Chaperonin 60; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D054303:HIV Reverse Transcriptase