Mutations in the amiA locus of the Gram-positive bacterium Streptococcus pneumoniae confer a complex phenotype including resistance to various antineoplastic drugs. As a first step towards the understanding of the molecular organization and the function(s) of this locus, we have cloned DNA fragments carrying its 5'- and 3'-extremities. We have isolated and characterized a down-promoter mutation and have located the functional limits of the locus. The amiA locus is between 5.8 and 7.5 kb long strongly suggesting that it encodes several proteins.