Koen Verhoef

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The current human immunodeficiency virus type 1 (HIV-1) shows an increasing number of distinct viral subtypes, as well as viruses that are recombinants of at least two subtypes. Although no biological differences have been described so far for viruses that belong to different subtypes, there is considerable sequence variation between the different HIV-1(More)
Human immunodeficiency viruses HIV-1 and HIV-2 encode a Tat protein that specifically activates transcription from the viral long terminal repeat. To characterize the properties of the Tat proteins, we have expressed them in Escherichia coli. The purified Tat protein was biochemically analyzed and tested for activity upon electroporation into human cell(More)
Live-attenuated human immunodeficiency virus type 1 (HIV-1) variants have shown great promise as AIDS vaccines, but continued replication can lead to the selection of faster-replicating variants that are pathogenic. We therefore designed HIV-1 genomes that replicate exclusively upon addition of the nontoxic effector doxycycline (dox). This was achieved by(More)
We present viral evolution as a novel and powerful method to optimize non-viral proteins. We used this approach to optimize the tetracycline (Tc)-regulated gene expression system (Tet system) for its function in mammalian cells. The components of the Tet system were incorporated in the human immunodeficiency virus (HIV)-1 virus such that viral replication(More)
The HIV-1 RNA genome forms dimers through base pairing of a palindromic 6-mer sequence that is exposed in the loop of the dimer initiation signal (DIS) hairpin structure (loop-loop kissing). The HIV-1 leader RNA can adopt a secondary structure conformation that is not able to dimerize because the DIS hairpin is not folded. Instead, this DIS motif is(More)
Human immunodeficiency virus type 1 (HIV-1) transcription is regulated by the viral Tat protein and cellular factors, of which the concentration and activity may depend on the cell type. Viral long terminal repeat (LTR) promoter sequences are therefore optimized to suit the specific nuclear environment of the target host cell. In long-term cultures of a(More)
Tat is an essential protein of human immunodeficiency virus type 1 (HIV-1) and activates transcription from the viral long terminal repeat (LTR) promoter. The tat gene is composed of two coding exons of which the first, corresponding to the N-terminal 72 amino acid residues, has been reported to be sufficient for its transcription function. We introduced a(More)
A major concern associated with the use of vaccines based on live-attenuated viruses is the possible and well documented reversion to pathogenic phenotypes. In the case of HIV, genomic deletions or mutations introduced to attenuate viral pathogenicity can be repaired by selection of compensating mutations. These events lead to increased virus replication(More)
The Tat protein of human immunodeficiency virus type 1 (HIV-1) is a potent trans-activator of transcription from the viral LTR promoter. Previous mutagenesis studies have identified domains within Tat responsible for binding to its TAR RNA target and for transcriptional activation. The minimal Tat activation domain is composed of the N-terminal 48 residues,(More)
We previously constructed a large set of mutants of the human immunodeficiency virus type 1 (HIV-1) regulatory protein Tat with conservative amino acid substitutions in the activation domain. These Tat variants were analyzed in the context of the infectious virus, and several mutants were found to be defective for replication. In an attempt to obtain(More)