The lysosomal compartment of human monocytic cells has never been investigated 
by a proteomic approach. By a combination of one-dimensional (1-D) and 
two-dimensional (2-D) gel electrophoresis, protein identification by N-terminal 
sequencing, matrix assisted laser desorption/ionization-mass spectrometry 
(MALDI-MS) peptide mass fingerprinting and tandem mass spectrometry (MS/MS) 
peptide sequence analysis, we initiated an exhaustive study of the human 
lyososomal proteome, which aims at establishing a 2-D reference map of human 
soluble lyososomal proteins. Human monocytic U937 cells were induced to secrete 
lysosomal soluble hydrolases by addition of NH4Cl in the culture medium. Since 
lysosomal soluble proteins are characterized by the presence of 
mannose-6-phosphate, they were purified on an affinity support bearing 
mannose-6-phosphate receptor. Analysis of the purified fraction led to the 
preliminary identification of fifteen proteins, among which twelve are 
well-known lysosomal hydrolases, one is assumed to be lysosomal on the basis of 
sequence homology to cysteine proteinases of the papain family, and two 
(leukocystatin and the human cellular repressor of E1A-stimulated genes) are 
described here for the first time as mannose-6-phosphate-containing proteins.