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Substrate specificity of human metallocarboxypeptidase D: Comparison of the two active carboxypeptidase domains

Fig 4

Quantitative peptidomics scheme for the characterization of rhCPD substrate specificity using the tryptic peptide library and representative spectra.

(A) Quantitative peptidomics scheme. Tryptic peptides were obtained from digestion of five selected proteins (BSA, bovine thyroglobulin, bovine α-lactalbumin and human α and β–hemoglobin) with trypsin. The resultant peptide library was aliquoted and digested with no enzyme or different rhCPD concentrations of 0.1, 1, 10, and 100 nM for 16 h at 37°C. After incubation samples were labeled with one of five stable isotopic TMAB-NHS tags (D0 = 100 nM; D3 = 10 nM; D6 = 1 nM; D9 = 0.1 nM; D12 = No enzyme). Then, samples were pooled and analyzed by LC-MS. Examples of representative data are shown for (B) non-substrates, (C) good substrates, (D) weak substrates and (E) products.

Fig 4