Cystatin F is a glycosylated human low molecular weight cysteine proteinase inhibitor.
A previously undescribed human member of the cystatin superfamily called cystatin F has been identified by expressed sequence tag sequencing in human cDNA libraries. A full-length cDNA clone was obtained from a library made from mRNA of CD34-depleted cord blood cells. The sequence of the cDNA contained an open reading frame encoding a putative 19-residue signal peptide and a mature protein of 126 amino acids with two disulfide bridges and enzyme-binding motifs homologous to those of Family 2 cystatins. Unlike other human cystatins, cystatin F has 2 additional Cys residues, indicating the presence of an extra disulfide bridge stabilizing the N-terminal region of the molecule. Recombinant cystatin F was produced in a baculovirus expression system and characterized. The mature recombinant protein processed by insect cells had an N-terminal segment 7 residues longer than that of cystatin C and displayed reversible inhibition of papain and cathepsin L (Ki = 1.1 and 0.31 nM, respectively), but not cathepsin B. Like cystatin E/M, cystatin F is a glycoprotein, carrying two N-linked carbohydrate chains at positions 36 and 88. An immunoassay for quantification of cystatin F showed that blood contains low levels of the inhibitor (0.9 ng/ml). Six B cell lines in culture secreted barely detectable amounts of cystatin F, but several T cell lines and especially one myeloid cell line secreted significant amounts of the inhibitor. Northern blot analysis revealed that the cystatin F gene is primarily expressed in peripheral blood cells and spleen. Tissue expression clearly different from that of the ubiquitous inhibitor, cystatin C, was also indicated by a high incidence of cystatin F clones in cDNA libraries from dendritic and T cells, but no clones identified by expressed sequence tag sequencing in several B cell libraries and in >600 libraries from other human tissues and cells.
Ni
J
J
Human Genome Sciences, Inc., Rockville, Maryland 20850-3338, USA.
Fernandez
M A
MA
Danielsson
L
L
Chillakuru
R A
RA
Zhang
J
J
Grubb
A
A
Su
J
J
Gentz
R
R
Abrahamson
M
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D014408:Biomarkers, Tumor; D003001:Cloning, Molecular; D055316:Cystatin C; D015891:Cystatins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D018076:DNA, Complementary; D016208:Databases, Factual; D015723:Gene Library; D006023:Glycoproteins / Q000737:chemistry; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010802:Phylogeny; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D014408:Biomarkers, Tumor; C525030:CST3 protein, human; C116891:CST7 protein, human; D055316:Cystatin C; D015891:Cystatins; D018076:DNA, Complementary; D006023:Glycoproteins; D011994:Recombinant Proteins