The Drosophila nuclear proteins Bj6 and Bx42 characterized previously are detected in a series of developmentally active puffs on salivary gland chromosomes. Here the binding of both proteins at puff 3C11-12 containing the glue protein gene Sgs-4 is described in more detail. By deletion analysis we show that both proteins bind within a chromosomal segment containing 17-19 kb of DNA surrounding the Sgs-4 gene. They are detectable at this site during the intermoult stages, before the puff regresses in response to the moulting hormone ecdysone. If the Sgs-4 gene together with flanking DNA sequences is brought into a different chromosomal position by P element transfer, both proteins are detected at this new location. Both proteins are bound to the chromosome within the range of 2.5 kb DNA upstream of the Sgs-4 gene. A strain containing a 52 bp deletion within this region fails to bind Bx42 protein suggesting that the missing DNA, which overlaps a hypersensitive region, may be required for the binding of the Bx42 protein.