The human pim-1 gene was recently identified as a new putative oncogene located on chromosome 6p21, a region showing karyotypic abnormalities in particular leukemias. In the present work we characterized the pim protein product. In vitro translation of positively selected poly(A)+ mRNA indicates that this gene encodes a 33-kilodalton protein. Anti-pim antibodies were raised against a fused TrpE-pim protein induced in a bacterial expression vector. This antibody immunoprecipitated a 33-kilodalton protein from in vivo [35S]methionine-labeled K562 and KCl myelogenous origin cell lines. This protein was localized to the cytoplasm, and in vivo labeling as well as in vitro kinase assay suggests that it is a phosphoprotein with tyrosine kinase activity. This was further confirmed by performing autophosphorylation directly on a p33pim-containing gel band cut out after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The results imply that the tyrosine kinase activity of pim can be recovered after boiling the pim-1 protein in sample buffer: a feature not described yet for this class of protein. These results suggest that pim-1 is a new member of the subgroup of oncogenes encoding tyrosine kinases.