The deaD gene of Klebsiella pneumoniae was isolated and its nucleotide sequence determined. The K. pneumoniae gene is highly homologous with the Escherichia coli analog throughout most of the coding region. The deduced primary sequence of the K. pneumoniae deaD gene product is 659 amino acids in length, in contrast with the 571 amino acids of the E. coli deaD product published previously. Sequence comparison revealed several differences near the 3' end of the deaD genes which result in the frame-shift effect. The 3' end sequence of the E. coli deaD gene was therefore analyzed to verify the discrepancy. Our result indicates that the E. coli deaD gene encodes a product of comparable size to the K. pneumoniae DeaD protein, and the carboxyl terminal sequences of the two proteins are highly homologous. In vivo expression of the K. pneumoniae deaD gene in E. coli yielded a 65-kDa protein. Primer extension analysis of the mRNA from K. pneumoniae identified a major transcription start site at an A residue 44 nt upstream of the first in-frame ATG codon.