A reversibly modified ‘hot start’ RNase H enzyme composition is described for the improved CATACLEAVE™ probe detection of nucleic acid sequences in a test sample. A key feature of the enzyme composition is the ability to regulate the catalytic activity of the RNase H during the course of a reverse transcription-PCR cycle. Thus, RNase H activity can be initially suppressed to minimize degradation of RNA:DNA primer heteroduplexes prior to reverse transcription. After cDNA synthesis is complete, RNase H activity is induced to promote the cleavage and fluorescent detection of CATACLEAVE™ probes that anneal to target DNA sequences within the reverse transcriptase-PCR products. The inducible RNase H enzyme is amenable to high throughput applications requiring one step reverse transcriptase CATACLEAVE™ PCR in a single reaction mix.