Nuclear factor kappaB (NF-kappaB) transcription factors regulate a large number of genes in response to inflammation, infection and stressful conditions. In this study, we investigated whether NF-kappaB p65 regulates the transcription of target genes by interacting with components of the basal transcription machinery. We examined the interaction of p65 with the basal transcription factor IIB (TFIIB). Glutathione S-transferase pull down assays showed that the Rel homology domain of p65 is important for binding to TFIIB. Molecular modelling, together with the generation of specific point mutants, revealed that residues 41 R and 42 S in the Rel homology domain of p65 facilitate the interaction with TFIIB. Mutation of these residues showed a decrease in p65 induced transcription, suggesting that they are involved in a functional interaction with TFIIB.