Peptidomic profiling of human cerebrospinal fluid identifies YPRPIHPA as a novel substrate for prolylcarboxypeptidase.

Prolylcarboxypeptidase (PRCP) is a serine protease that catalyzes the cleavage of C-terminal amino acids linked to proline in peptides. It is ubiquitously expressed and is involved in regulating blood pressure, proliferation, inflammation, angiogenesis, and weight maintenance. To identify the candidate proximal target engagement markers for PRCP inhibition in the central nervous system, we profiled the peptidome of human cerebrospinal fluid to look for PRCP substrates using a MS-based in vitro substrate profiling assay. These experiments identified a single peptide, with the sequence YPRPIHPA, as a novel substrate for PRCP in human cerebrospinal fluid. The peptide YPRPIHPA is from the extracellular portion of human endothelin B receptor-like protein 2.

Expression of human brain carboxypeptidase B, a possible cleaving enzyme for beta-amyloid precursor protein, in peripheral fluids.

Human brain carboxypeptidase B (HBCPB) is a novel brain protease that processes native brain beta-amyloid precursor protein (APP) in vitro. Immunoblot analysis of human serum and cerebrospinal fluid (CSF) using anti C14-module antibody, which recognizes the C-terminal peptide unique to HBCPB, detected the 30 and 40 kDa immunoreactive bands. Analysis of HBCPB prepared from both serum and CSF demonstrated proteolytic activities for brain APP. Protease inhibitor spectrum analysis also supports that these bands correspond to the mature form and and prepro form of HBCPB, respectively. As is the case in brain parenchyma, the prepro-form is dominant in CSF. In serum, however, the majority of HBCPB exists in the mature form, possibly due to an abundant trypsin-like proteolytic activity in serum. HBCPB expressed in serum and CSF, therefore, may have a significance as a peripheral marker of the brain protease, which participates in APP processing in human brain.