HIV RT plays a key role in viral replication and is an important target for development of anti-HIV drugs. However, the turn between helices H and I of HIV-1 RT thumb region (amino acid residues 269 to 277) has not been well characterized yet. To understand the structure-function relationship of this turn in details, we investigated whether alanine substitutions in this region would affect RT activity. Except for residues 269 and 270, which have been reported to have no significant influence on RT activity [9, 10] , and for residue 272, which is originally an alanine, six mutant pseudoviruses (residues 271 and 273 to 277) were constructed and viral replication was compared with that of the wild type virus. The replication of two mutants, Y271A and I271A, were found to be almost completely abolished (Fig. 1A) . This result was further confirmed in live virus system (Fig. 1B &  1C) . Since it has been reported that bacterially expressed Y271A mutant has only 1% activity of wild type enzyme [9] , we asked whether the mutants would similarly affect RT activity in the viral particles. The results showed that the RT activity was basically undetectable in viral particles of these two mutants ( Fig. 2A) . It was also found that the loss of RT activity in the viral particles might be attributed to the absence of RT in the viral particles rather than the incorporation of dysfunctional RT into the virions (Fig. 2B) . After viral entry into the cells, the first step of HIV-1 replication is reverse transcription. Our results thus suggested that replication of mutant viruses was abrogated, because the mutant RT did not exist in the virion.