Human cathepsin F. Molecular cloning, functional expression, tissue localization, and enzymatic characterization.
A cDNA for a novel human papain-like cysteine protease, designated cathepsin F, has been cloned from a lambdagt10-skeletal muscle cDNA library. The nucleotide sequence encoded a polypeptide of 302 amino acids composed of an 88-residue propeptide and a 214-residue mature protein. Protein sequence comparisons revealed 58% homology with cathepsin W; about 42-43% with cathepsins L, K, S, H, and O; and 38% with cathepsin B. Sequence comparisons of the propeptides indicated that cathepsin F and cathepsin W may form a new cathepsin subgroup. Northern blot analysis showed high expression levels in heart, skeletal muscle, brain, testis, and ovary; moderate levels in prostate, placenta, liver, and colon; and no detectable expression in peripheral leukocytes and thymus. The precursor polypeptide of human recombinant cathepsin F, produced in Pichia pastoris, was processed to its active mature form autocatalytically or by incubation with pepsin. Mature cathepsin F was highly active with comparable specific activities toward synthetic substrates as reported for cathepsin L. The protease had a broad pH optimum between 5.2 and 6.8. Similar to cathepsin L, its pH stability at cytosolic pH (7.2) was short, with a half-life of approximately 2 min. This may suggest a function in an acidic cellular compartment. Transient expression of T7-tagged cathepsin F in COS-7 cells revealed a vesicular distribution of the gene product in the juxtanuclear region of the cells. However, contrary to all known cathepsins, the open reading frame of the cathepsin F cDNA did not encode a signal sequence, thus suggesting that the protease is targeted to the lysosomal compartment via an N-terminal signal peptide-independent lysosomal targeting pathway.
Wang
B
B
Department of Human Genetics, Mount Sinai School of Medicine, CUNY, New York, New York 10029, USA.
Shi
G P
GP
Yao
P M
PM
Li
Z
Z
Chapman
H A
HA
Brömme
D
D
D000595:Amino Acid Sequence; D001483:Base Sequence; D056646:Cathepsin F; D056664:Cathepsin W; D002403:Cathepsins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases / Q000737:chemistry; D018076:DNA, Complementary; D005260:Female; D015723:Gene Library; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008297:Male; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000201:enzymology*; D009928:Organ Specificity; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D015183:Restriction Mapping; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014158:Transcription, Genetic
D018076:DNA, Complementary; D011994:Recombinant Proteins; D002403:Cathepsins; C534187:CTSW protein, human; D056664:Cathepsin W; D003546:Cysteine Endopeptidases; C534090:CTSF protein, human; D056646:Cathepsin F