Previous studies have demonstrated that NE, PR3, and CG colocalize with HNPs in the azurophil granules of neutrophils. Kamdar et al. demonstrated that NE digestion of proRMAD4, precursor of rhesus myeloid a-defensin-4 (RMAD4), converted proRMAD4 to mature RMAD4. However, digestion of pro-RMAD4 with PR3 and CG generated RMAD4 variants with Nterminal extensions (Fig. 5D ) [32] . To test for the role of these granule serine proteases in human HNP processing, proHNP1A was incubated in separate reactions with NE, PR3, or CG, and the RP-HPLC-purified cleavage products were analyzed by MALDI-TOF MS and analytical HPLC ( Fig. 5 and Table 1 ). Incubation of His 6 -proHNP1 with NE or PR3 generated peptide products that coeluted with native HNP1 in RP-HPLC (34. Table 1 ). The identities of HNP1 derivatives were also confirmed by MALDI-TOF MS of each peptide following reduction and S-carboxyamidomethylation. In each case reduction and alkylation produced a mass increase of 348 A.M.U., confirming the presence of 6 disulfide-linked cysteine residues ( Table 1 ). The NE-and PR3-derived peptides also comigrated with native HNP1 on AU-gels ( Fig. 5C ; lanes 1-3). Thus, NE and PR3 processing of His 6 -proHNP1 generated a peptide that was indistinguishable from natural HNP1.