The Olfactory Specific-E and -F genes (OS-E and OS-F) belong to the odorant-binding protein gene family, which includes the general odorant-binding proteins and the pheromone-binding proteins. In Drosophila melanogaster, these genes are arranged in tandem in a genomic region near the centromere of chromosome arm 3R. We examined the pattern of DNA sequence variation in an approximately 7-kb genomic region encompassing the two OS genes in four species of the melanogaster subgroup of Drosophila and in a population sample of D. melanogaster. We found that both the OS-E and the OS-F gene are present in all surveyed species. Nucleotide divergence estimates would support that the two genes are functional, although they diverge in their functional constraint. The pattern of nucleotide variation in D. melanogaster also differed between genes. Variation in the OS-E gene region exhibited an unusual and distinctive pattern: (i) a relatively high number of fixed amino acid replacements in the encoded protein and (ii) a peak of nucleotide polymorphism around the OS-E gene. These results are unlikely under the neutral model and suggest the action of natural selection in the evolution of the two odorant-binding protein genes.