title: Identification of residues in the C-terminal domain of HIV-1 integrase that mediate binding to the transportin-SR2 protein.
abstract: Transportin-SR2 (TRN-SR2 and TNPO3) is a cellular cofactor of HIV replication that has been implicated in the nuclear import of HIV. TRN-SR2 was originally identified in a yeast two-hybrid screen as an interaction partner of HIV integrase (IN) and in two independent siRNA screens as a cofactor of viral replication. We have now studied the interaction of TRN-SR2 and HIV IN in molecular detail and identified the TRN-SR2 interacting regions of IN. A weak interaction with the catalytic core domain (CCD) and a strong interaction with the C-terminal domain (CTD) of IN were detected. By dissecting the catalytic core domain (CCD) of IN into short structural fragments, we identified a peptide (INIP(1), amino acids (170)EHLKTAVQMAVFIHNFKRKGGI(191)) retaining the ability to interact with TRN-SR2. By dissecting the C-terminal domain (CTD) of IN, we could identify two interacting peptides (amino acids (214)QKQITKIQNFRVYYR(228) and (262)RRKVKIIRDYGK(273)) that come together in the CTD tertiary structure to form an exposed antiparallel β-sheet. Through site-specific mutagenesis, we defined the following sets of amino acids in IN as important for the interaction with TRN-SR2: Phe-185/Lys-186/Arg-187/Lys-188 in the CCD and Arg-262/Arg-263/Lys-264 and Lys-266/Arg-269 in the CTD. An HIV-1 strain carrying K266A/R269A in IN was replication-defective due to a block in reverse transcription, confounding the study of nuclear import. Insight into the IN/TRN-SR2 interaction interface is necessary to guide drug discovery efforts targeting the nuclear entry step of replication.
authors.affiliation: Laboratory for Molecular Virology and Gene Therapy, Katholieke Universiteit Leuven, 3000 Leuven, Belgium.
authors.forename: Stephanie
authors.initials: S
authors.lastname: De Houwer
authors.forename: Jonas
authors.initials: J
authors.lastname: Demeulemeester
authors.forename: Wannes
authors.initials: W
authors.lastname: Thys
authors.forename: Oliver
authors.initials: O
authors.lastname: Taltynov
authors.forename: Katarina
authors.initials: K
authors.lastname: Zmajkovicova
authors.forename: Frauke
authors.initials: F
authors.lastname: Christ
authors.forename: Zeger
authors.initials: Z
authors.lastname: Debyser
mesh_terms: D019943:Amino Acid Substitution; D019427:HIV Integrase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D010449:Peptide Mapping; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D048348:Reverse Transcription / Q000502:physiology; D028961:beta Karyopherins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D010455:Peptides; C546476:TNPO3 protein, human; D028961:beta Karyopherins; D019427:HIV Integrase