scholarly journals Correlation between Viral Resistance to Zidovudine and Resistance at the Reverse Transcriptase Level for a Panel of Human Immunodeficiency Virus Type 1 Mutants

2001 ◽  
Vol 75 (15) ◽  
pp. 7202-7205 ◽  
Author(s):  
Johan Lennerstrand ◽  
Kurt Hertogs ◽  
David K. Stammers ◽  
Brendan A. Larder
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Immunodeficiency Virus ◽  
Azt Resistance ◽  
Resistance Data ◽  
Insertion Mutations ◽  
Dependent Mechanism

ABSTRACT Using a large panel of human immunodeficiency virus type 1 site-directed mutants, we have observed a higher correlation than has previously been demonstrated between zidovudine (AZT)-triphosphate resistance data at the reverse transcriptase (RT) level and corresponding viral AZT resistance. This enhanced-resistance effect at the RT level was seen with ATP and to a lesser extent with PPi when ATP was added at physiological concentrations. The ATP-dependent mechanism (analogous to pyrophosphorolysis) appears to be dominant in the mutants bearing the D67N and K70R or 69 insertion mutations, whereas the Q151M mutation seems independent of ATP for decreased binding to AZT-triphosphate.

scholarly journals The M184V Mutation Reduces the Selective Excision of Zidovudine 5′-Monophosphate (AZTMP) by the Reverse Transcriptase of Human Immunodeficiency Virus Type 1

2002 ◽  
Vol 76 (7) ◽  
pp. 3248-3256 ◽  
Author(s):  
Paul L. Boyer ◽  
Stefan G. Sarafianos ◽  
Edward Arnold ◽  
Stephen H. Hughes
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Published Data ◽  
Immunodeficiency Virus ◽  
Azt Resistance ◽  
Hiv 1 ◽  
M184v Mutation

ABSTRACT The M184V mutation in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) causes resistance to lamivudine, but it also increases the sensitivity of the virus to zidovudine (3′-azido-3′-deoxythymidine; AZT). This sensitization to AZT is seen both in the presence and the absence of the mutations that confer resistance to AZT. AZT resistance is due to enhanced excision of AZT 5′-monophosphate (AZTMP) from the end of the primer by the RT of the resistant virus. Published data suggest that the excision reaction involves pyrophosphorolysis but that the likely in vivo pyrophosphate donor is not pyrophosphate but ATP. The mutations that lead to AZT resistance enhance ATP binding and, in so doing, enhance pyrophosphorolysis. The excision reaction is specific for AZT because HIV-1 RT, which can form a closed complex with a dideoxy-terminated primer and an incoming deoxynucleoside triphosphate (dNTP), does not form the closed complex with an AZTMP-terminated primer and an incoming dNTP. This means that an AZTMP-terminated primer has better access to the site where it can be excised. The M184V mutation alters the polymerase active site in a fashion that specifically interferes with ATP-mediated excision of AZTMP from the end of the primer strand. The M184V mutation does not affect the incorporation of AZT 5′-triphosphate (AZTTP), either in the presence or the absence of mutations that enhance AZTMP excision. However, in the presence of ATP, the M184V mutation does decrease the ability of HIV-1 RT to carry out AZTMP excision. Based on these results, and on the results of other excision experiments, we present a model to explain how the M184V mutation affects AZTMP excision.

scholarly journals High-Level Resistance to 3′-Azido-3′-Deoxythimidine due to a Deletion in the Reverse Transcriptase Gene of Human Immunodeficiency Virus Type 1

2000 ◽  
Vol 74 (2) ◽  
pp. 1023-1028 ◽  
Author(s):  
Tomozumi Imamichi ◽  
Tanima Sinha ◽  
Hiromi Imamichi ◽  
Yi-Ming Zhang ◽  
Julie A. Metcalf ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Antiretroviral Therapy ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Immunodeficiency Virus ◽  
Azt Resistance ◽  
High Level ◽  
Level Resistance

ABSTRACT A variant of human immunodeficiency virus type 1 (HIV-1) possessing a deletion in the reverse transcriptase (RT) gene at codon 67 was identified in a patient who had failed combination antiretroviral therapy. This deletion initially emerged under the selective pressure of combination therapy with 3′-azido-3′-deoxythymidine (AZT) plus 2′,3′-dideoxyinosine. It has persisted for more than 3 years in association with the accumulation of a variety of other well-described drug resistance mutations and an uncharacterized mutation at RT codon 69 (T69G). Phenotypic studies demonstrated that the codon 67 deletion by itself had little effect on AZT sensitivity. However, in the context of the T69G mutation and three other mutations known to be associated with AZT resistance (K70R, T215F, and K219Q), this deletion led to a increase in AZT resistance from 8.5-fold to 445-fold. A further increase in resistance (up to 1,813-fold) was observed when two mutations associated with nonnucleoside RT inhibitor resistance (K103N and L74I) were added to the deletion T69G K70R T215F K219Q construct. Hence, these results establish that a deletion at RT codon 67 may be selected for in the presence of antiretroviral therapy and may lead to high-level resistance to AZT.

scholarly journals Effects of Dipeptide Insertions between Codons 69 and 70 of Human Immunodeficiency Virus Type 1 Reverse Transcriptase on Primer Unblocking, Deoxynucleoside Triphosphate Inhibition, and DNA Chain Elongation

2003 ◽  
Vol 77 (6) ◽  
pp. 3871-3877 ◽  
Author(s):  
Peter R. Meyer ◽  
Johan Lennerstrand ◽  
Suzanne E. Matsuura ◽  
Brendan A. Larder ◽  
Walter A. Scott
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Chain Elongation ◽  
Immunodeficiency Virus ◽  
Deoxynucleoside Triphosphate ◽  
Dna Chain ◽  
Insertion Mutations

ABSTRACT Finger insertion mutations of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) (T69S mutations followed by various dipeptide insertions) have a multinucleoside resistance phenotype that can be explained by decreased sensitivity to deoxynucleoside triphosphate (dNTP) inhibition of the nucleotide-dependent unblocking activity of RT. We show that RTs with SG or AG (but not SS) insertions have three- to fourfold-increased unblocking activity and that all three finger insertion mutations have threefold-decreased sensitivity to dNTP inhibition. The additional presence of M41L and T215Y mutations increased unblocking activity for all three insertions, greatly reduced the sensitivity to dNTP inhibition, and resulted in defects in in vitro DNA chain elongation. The DNA chain elongation defects were partially repaired by additional mutations at positions 210, 211, and 214. These results suggest that structural communication between the regions of RT defined by these mutations plays a role in the multinucleoside resistance phenotype.

scholarly journals Relationship between 3′-Azido-3′-Deoxythymidine Resistance and Primer Unblocking Activity in Foscarnet-Resistant Mutants of Human Immunodeficiency Virus Type 1 Reverse Transcriptase

2003 ◽  
Vol 77 (11) ◽  
pp. 6127-6137 ◽  
Author(s):  
Peter R. Meyer ◽  
Suzanne E. Matsuura ◽  
Dianna Zonarich ◽  
Rahul R. Chopra ◽  
Eric Pendarvis ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Binding Site ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Resistance Mutations ◽  
Immunodeficiency Virus ◽  
Azt Resistance ◽  
Hiv 1

ABSTRACT Phosphonoformate (foscarnet) is a pyrophosphate (PPi) analogue and a potent inhibitor of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT), acting through the PPi binding site on the enzyme. HIV-1 RT can unblock a chain-terminated DNA primer by phosphorolytic transfer of the terminal residue to an acceptor substrate (PPi or a nucleotide such as ATP) which also interacts with the PPi binding site. Primer-unblocking activity is increased in mutants of HIV-1 that are resistant to the chain-terminating nucleoside inhibitor 3′-azido-3′-deoxythymidine (AZT). We have compared the primer-unblocking activity for HIV-1 RT containing various foscarnet resistance mutations (K65R, W88G, W88S, E89K, S117T, Q161L, M164I, and the double mutant Q161L/H208Y) alone or in combination with AZT resistance mutations. The level of primer-unblocking activity varied over a 150-fold range for these enzymes and was inversely correlated with foscarnet resistance and directly correlated with AZT resistance. Based on published crystal structures of HIV-1 RT, many of the foscarnet resistance mutations affect residues that do not make direct contact with the catalytic residues of RT, the incoming deoxynucleoside triphosphate (dNTP), or the primer-template. These mutations may confer foscarnet resistance and reduce primer unblocking by indirectly decreasing the binding and retention of foscarnet, PPi, and ATP. Alternatively, the binding position or orientation of PPi, ATP, or the primer-template may be changed in the mutant enzyme complex so that molecular interactions required for the unblocking reaction are impaired while dNTP binding and incorporation are not.

scholarly journals Effects of Specific Zidovudine Resistance Mutations and Substrate Structure on Nucleotide-Dependent Primer Unblocking by Human Immunodeficiency Virus Type 1 Reverse Transcriptase

2002 ◽  
Vol 46 (5) ◽  
pp. 1540-1545 ◽  
Author(s):  
Peter R. Meyer ◽  
Suzanne E. Matsuura ◽  
Adviye A. Tolun ◽  
Ingrid Pfeifer ◽  
Antero G. So ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Resistance Mutations ◽  
Immunodeficiency Virus ◽  
Azt Resistance ◽  
The Difference

ABSTRACT Nucleotide-dependent unblocking of chain-terminated DNA by human immunodeficiency virus type 1 reverse transcriptase (RT) is enhanced by the presence of mutations associated with 3′-azido-3′-deoxythymidine (AZT) resistance. The increase in unblocking activity was greater for mutant combinations associated with higher levels of in vivo AZT resistance. The difference between mutant and wild-type activity was further enhanced by introduction of a methyl group into the nucleotide substrate and was decreased for a nonaromatic substrate, suggesting that π-π interactions between RT and an aromatic structure may be facilitated by these mutations.

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