A
cDNA clone encoding a human NK
serine protease was obtained by screening a lambda-gt10 library from the Lopez NK
leukemia with the rat natural killer Met-ase (RNK-Met-1)
cDNA clone. In Northern blot analysis human Met-ase (Hu-Met-1)
cDNA hybridized with a 0.9-kb
mRNA in two human NK
leukemia cell lines, unstimulated human PBMC, and untreated purified CD3-CD56+ large granular lymphocytes. Unlike other members of the
granzyme family that are highly expressed in activated peripheral T cells, the
Hu-Met-1 transcript was barely detected in a population of PMA and
ionomycin or IL-2-treated high density T cells. Several in vitro cultured Burkitt
lymphomas, chronic- and promyeloid
leukemias, acute lymphoblastic
leukemias, and colon and ovarian
carcinomas and colon and ovarian
carcinomas did not express
Hu-Met-1 mRNA.
Hu-Met-1 mRNA expression in a small number of human T cell
tumor lines did not correlate with any particular phenotype or stage of development. The presence of
Hu-Met-1 mRNA closely correlated with the Met-ase activity of cellular lysates prepared from these various human peripheral blood subsets and in vitro cultured cell lines. Met-ase activity detected in whole cell lysates of cytotoxic lymphocytes was associated with the cytoplasmic granules of these cells. The nucleotide sequence of the
Hu-Met-1 cDNA clone encodes a predicted
serine protease of 257
amino acids. The predicted
protein is an active
enzyme of 232
amino acids with a calculated unglycosylated m.w. of 27,100.
Hu-Met-1 is 66% identical to
RNK-Met-1 at the
amino acid level. The human and rat mature
protein sequences conserve the active site His, Asp, and Ser
amino acids that form the catalytic triad of
serine proteases, all 8
cysteine residues, and several
amino acids critical in the formation of the substrate binding pocket. The gene for the
Hu-Met-1 serine protease is located on chromosome 19, which distinguishes it from any other member of the human
granzyme family.