Characterization of the functional domains of human foamy virus integrase using chimeric integrases.
Retroviral integrases insert viral DNA into target DNA. In this process they recognize their own DNA specifically via functional domains. In order to analyze these functional domains, we constructed six chimeric integrases by swapping domains between HIV-1 and HFV integrases, and two point mutants of HFV integrase. Chimeric integrases with the central domain of HIV-1 integrase had strand transfer and disintegration activities, in agreement with the idea that the central domain determines viral DNA specificity and has catalytic activity. On the other hand, chimeric integrases with the central domain of HFV integrase did not have any enzymatic activity apart from FFH that had weak disintegration activity, suggesting that the central domain of HFV integrase was defective catalytically or structurally. However, these inactive chimeras were efficiently complemented by the point mutants (D164A and E200A) of HFV integrase, indicating that the central domain of HFV integrase possesses potential enzymatic activity but is not able to recognize viral or target DNA without the help of its homologous N-terminal and C-terminal domains.
Lee
Hak Sung
HS
Department of Biotechnology and BET Research Institute, Chung-Ang University, Ansung 456-756, Korea.
Kang
Seung Yi
SY
Shin
Cha-Gyun
CG
D000595:Amino Acid Sequence; D002384:Catalysis; D004279:DNA, Viral / Q000378:metabolism; D019427:HIV Integrase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D019426:Integrases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D016092:Spumavirus / Q000201:enzymology*; D016662:Virus Integration
D004279:DNA, Viral; D011993:Recombinant Fusion Proteins; D019427:HIV Integrase; D019426:Integrases