Xanthomonas campestris pv. campestris produces three proteases, Prt1, Prt2, and Prt3, the first two of which are involved in pathogenicity. In this study, nucleotide A 84 nt upstream of the prt1 start codon, which is 8 nt downstream of the -10 sequence, was determined as the transcription start site by the 5' RACE (rapid amplification of cDNA ends) method. Using Pprt1-lacZ transcriptional fusion constructs for assays, several interesting characteristics of prt1 promoter were revealed. The expression is inducible by LB medium or casein proteins and involves the global transcription factor Clp (cyclic AMP receptor protein-like protein). The region containing by -392 to -80 relative to the prt1 translation initiation codon is required for maximal expression, in which by -392 to -207 responds to the Clp-mediated regulation and the induction. In presence of inducers and the clp wild-type background, the levels of expression continue to increase following cell growth until 30h after the cultures entering stationary phase. Since prt1 promoter shows no response to stressful conditions and neither growth nor cell viability is affected by prt1 mutation, Prt1 appears to be a secondary metabolite of X. campestris pv. campestris. (C) 2002 Elsevier Science (USA). All rights reserved.